Assessing the analytical reliability of platelet counts and related indices in dogs and cats: a correlation matrix analysis comparing K2-EDTA and MgSO4 anticoagulation

Assessing the analytical reliability of platelet counts and related indices in dogs and cats: a correlation matrix analysis comparing K2-EDTA and MgSO4 anticoagulation | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Assessing the analytical reliability of platelet counts and related indices in dogs and cats: a correlation matrix analysis comparing K2-EDTA and MgSO4 anticoagulation Amir Hossein Tofighi Soleimandarabi, Helia Ghasemzadeh, Mahmood Ahmadi-hamedani, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8282197/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract The standard anticoagulant, ethylene diamine tetra-acetic acid (EDTA), often produces inaccurate platelet results (platelet count (PLT), plateletcrit (PCT), mean platelet volume (MPV), and platelet distribution width (PDW) in dog and cat blood due to its tendency to induce aggregation. This research, therefore, proposes magnesium sulfate (MgSO4) as the superior alternative because it actively prevents this clumping. The research examined blood samples from 31 feline and 32 canine individuals, totaling 122 blood tubes. Two types of tubes were used: k2-EDTA and MgSO4. Clinically healthy dogs and cats were used to obtain the samples, and PLT levels and related indices, such as MPV, PCT, and PDW, were measured using the Celltac α VET MEK-6550 hematology analyzer. In dogs, platelet parameters in MgSO4 and EDTA showed strong agreement ( r ≥ 0.90 for MPV, PLT, PCT, and PDW), with stability in MPV but a decrease in mean PCT in MgSO4 (0.30 vs. 0.56 in K2-EDTA). Feline results confirmed a strong correlation between PLT and PCT ( r = 0.99), but analytical agreement between K2-EDTA and MgSO4 was poor for MPV ( r = -0.11) and PDW ( r = 0.21). MgSO4 is effective as a replacement for K2-EDTA in dogs. MgSO4 in cats is critical because it helps prevent platelet clumping, thereby maintaining normal PLT and PCT levels. The beneficial compound in feline blood analysis requires caution in use. There are no favorable MgSO4 values for K2-EDTA with respect to the size parameters MPV and PDW. It suggests such parameters are not consistently accurate, irrespective of the anticoagulant used. Platelet Counts EDTA MgSO4 Dogs Cats Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction Accurately measuring platelet parameters, such as platelet count (PLT), mean platelet volume (MPV), plateletcrit (PCT), and platelet distribution width (PDW), is essential for evaluating coagulation, inflammation, and thrombotic risk in veterinary medicine. It is commonly accepted that in hematology laboratories, ethylene diamine tetra-acetic acid (EDTA) becomes the preferred anticoagulant due to its ability to chelate calcium and, consequently, inhibit coagulation development [ 2 , 14 ]. However, the use of EDTA can also lead to pre-analytical issues that invalidate test results. In humans, a rare side effect of EDTA is pseudothrombocytopenia (PTCP), in which the platelet count appears low because platelets clump together in the test tube. It occurs in about 0.07–0.27% of EDTA samples and is most often observed when samples are kept at room temperature [ 9 ]. Platelet aggregation is also a characteristic feature of canine and feline blood samples that can lead to misleading diagnoses of thrombocytopenia and to the inappropriate completion of diagnostic profiles or the supposed treatment process [ 11 , 23 ]. Cold agglutinin antibodies, autoantibody precipitation upon addition of EDTA, or low anticoagulant levels or platelet activation can result in platelet aggregation in a venous blood sample [ 20 , 22 ]. The development and progression of this process can differ among different species. It is less observed in dogs than in cats. Aggregation by platelets would be more common in felines, and the process could be found out easily due to the unique properties of feline platelets, such as their osmotic influence of EDTA and relatively smaller size [ 12 , 19 ]. This clumping may lead to inaccurate platelet counts, slowing the diagnostic test for thrombocytopenia. EDTA may also alter the cell's osmotic pressure, causing swelling that can make platelet volume measurements, such as MPV, appear higher than they really are [ 9 ]. Some studies in humans have shown that EDTA-PTCP is associated with IgG autoantibodies targeting a hidden region of the IIb unit of the GPIIbIIIa receptor (also known as CD41/CD61), which is revealed only in the presence of EDTA [ 2 ]. Researchers use three main ways to reduce the effects of PTCP in dogs and cats. The first is to prewarm refrigerated samples before analysis. The second is to add PLT antagonists, such as prostaglandin E1 (PGE1), citrate, theophylline, adenosine, dipyridamole (CTAD), and Iloprost, to collection tubes before drawing blood [ 19 , 21 ]. The third intervention involves pretreating EDTA tubes with amikacin, which neutralizes the fibrinogen receptor GPIIbIIIa and reduces its fibrinogen binding [ 20 ]. To reduce these pre-analytical problems, especially in preparations that are likely to exhibit platelet aggregation, other anticoagulants, including magnesium sulfate (MgSO4), are also added. MgSO4 works as an anticoagulant because magnesium ions compete with calcium in coagulation reactions and help inhibit clotting [ 4 ]. More importantly, MgSO4 is also recognized as an antiplatelet aggregation agent; magnesium, being a calcium antagonist, can inhibit platelet activation and prevent clumping [ 3 ]. Studies in people have shown that MgSO4 improves PLT function in those with PTCP. It means MgSO4 can give more true results [ 3 , 16 ]. Right now, there is no other choice like MgSO4 for EDTA-PTCP that can deliver better or more accurate results for dogs and cats. Given the species-specific challenges and the use of the specific hematology analyzer, Celtac-Alpha (Nihon Kohden), in our study, the main objective of this research is to compare the discrepancies and statistical correlations of PLT and related platelet indices between K2-EDTA and MgSO4 anticoagulants in dog and cat blood samples. Material and methods Studied animals and sample collection We conducted a cross-sectional study utilizing dogs and cats that presented for routine health check-ups at a private veterinary hospital in Tehran, Iran. A total of 61 client-owned dogs and cats (30 dogs and 31 cats) were ultimately selected for this study. All animals included in the study were clinically healthy, with no history of coagulation disorders, underlying platelet diseases, or use of anticoagulants or nonsteroidal anti-inflammatory drugs (NSAIDs). For each dog and cat included in the study, essential demographic data such as age, sex, breed, and the individual's name were carefully recorded along with the blood sample. No animals were euthanized or sacrificed for this study. Blood samples were collected from live animals as non-invasive clinical specimens, with no anesthesia required; the animals remained fully conscious throughout the collection process. Anticoagulants used For this study, we collected paired, simultaneous blood samples from 31 cats and 30 dogs (a total of 122 tubes) immediately after venipuncture. We used two types of tubes: K2-EDTA (S-Monovette K2-EDTA; fill volume 2.6 mL) as the routine standard for hematology, and MgSO4 (S-Monovette ThromboExact; fill volume 2.7 mL) [10]. The MgSO4 collection tubes were particularly selected as an anticoagulant method alternative based on research work that proposes it assists in combating PTCP, which stands for the phenomenon wherein the clotting agent EDTA causes the platelet cell masses to aggregate, thus producing an inflated cell count [3, 16]. Pre-analytical phase To prevent platelet preactivation, the blood was fully extended after the tourniquet was released, gently mixed with the anticoagulant, and immediately brought to the laboratory for further processing [10]. Platelet-related measurements, particularly sample storage, may result in volume changes in K2-EDTA. Aggregation, a property seen with feline platelets, would be enhanced [19]. To prevent unwanted changes, all hematological analyses were performed within 30 minutes of blood collection [9]. Measurement of platelet parameters PLT and platelet indices, including MPV, PCT, and PDW, were measured using the Celltac α VET MEK-6550 hematology analyzer (Nihon Kohden). This device operates using the impedance method and can accurately measure these parameters in canine and feline blood [9]. PCT was calculated as a function of PLT and MPV. Data analysis Measurements for each parameter were recorded in paired sets using both the K2-EDTA and MgSO4 anticoagulants to enable direct comparison. Initially, the Shapiro-Wilk test was performed on each parameter to assess its data distribution and normality. Since the majority of parameters (PLT, MPV, and PCT) did not exhibit a normal distribution in either dog or cat samples, the Wilcoxon Signed-Rank Test, a non-parametric method, was strategically employed to compare the effects of K2-EDTA versus MgSO4 on these values. Conversely, because the PDW parameter did follow a normal distribution, its mean values were compared using the Paired Samples t-Test, a parametric test. To evaluate the level of agreement between the two anticoagulants and to determine the relationships among all measured parameters, a Pearson’s Correlation Coefficient matrix was calculated. A strong correlation, close to 1, was considered representative of strong agreement between variables, implying the possibility of interchangeability. Data analysis and graphics processing were carried out in R-Studio (version 4.4.2). A p-value less than 0.05 was considered statistically significant. Results The platelet parameters, such as PLT, MPV, PCT, and PDW, were evaluated in whole-blood samples from 30 dogs and 31 cats using two anticoagulants: K2-EDTA (E) and MgSO4 (S). The exact statistical distribution, minimum, Q1, median, Q3, maximum, and mean, is given for each parameter in Tables 1 and 2. Comparison of PLT between K2-EDTA and MgSO4 anticoagulants in dogs and cats In the PLT range in K2-EDTA, from 177 to 956, the mean was 404.43, with a median of 394.0; in MgSO4, the range was similar, from 183 to 967, but the mean was slightly higher, at 418.63, with a median of 386.0. Such an increase in mean could indicate reduced platelet aggregation in MgSO4 (Table 1 and Fig. 1a). This parameter varied more in cats: in K2-EDTA, it ranged from 139 to 1232 (mean 485.16, median 369.00), whereas in MgSO4, the mean increased to 545.52 (range 53 to 1250, median 445.00). This difference of about 12% may be important for improving diagnostic accuracy in cats, where PLT is generally variable due to aggregation (Table 2 and Fig. 1b). Comparison of MPV between K2-EDTA and MgSO4 anticoagulants in dogs and cats In dogs, MPV was found to be highly stable, with a range of 5.60–15.10 in K2-EDTA (mean: 7.69, median: 7.30) and a comparably stable range in MgSO4 (mean: 7.64, median: 7.40), thus highlighting the MgSO4 anticoagulant’s strong capacity for maintaining platelet morphology (Table 1, Fig. 2a). In the case of cats, MPV was higher, with a range of 8.00–12.00 in K2-EDTA (mean: 9.78, median: 9.70) and 7.60 to 12.20 in MgSO4 (mean 9.50, median 9.30). This slight difference could indicate species-specific effects of anticoagulants on platelet volume, underscoring the importance of adjusting laboratory protocols (Table 2 and Fig. 2b). Comparison of PCT between K2-EDTA and MgSO4 anticoagulants in dogs and cats In dogs, PCT ranged from 0.14 to 0.73 in K2-EDTA with a mean of 0.56 and a median of 0.28, while in MgSO4, the mean decreased to 0.30 (range 0.11 to 0.72, median 0.27). This significant difference might reflect differences in the overall effect on platelet volume (Table 1 and Fig. 3a). In cats, this parameter increased significantly more in MgSO4; in K2-EDTA, the range was 0.12 to 1.27, with an average of 0.48 and a median of 0.37, while in MgSO4, the values ranged from 0.05 to 1.31, with an average value of 0.56 and a median of 0.42. An increase of 17% may represent one of the main discoveries for improved diagnostic performance in cats, underscoring the potential of MgSO4 to reduce variability (Table 2 and Fig. 3b). Comparison of PDW between K2-EDTA and MgSO4 anticoagulants in dogs and cats PDW in dogs was very similar, with a range of 14.10 to 17.30 in both anticoagulants (mean approximately 15.8 in both; median around 15.7), strongly emphasizing the reliability of MgSO₄ in measuring platelet size distribution (Table 1 and Fig. 4a). In cats, there was a larger spread: the range in EDTA was from 10.70 to 17.40 (mean 14.39, median 14.40), while in MgSO₄, it ranged from 8.90 to 21.60 (mean 15.49, median 15.70). The approximately 8% increase in mean platelet size suggests greater variation in the platelet size distribution under MgSO₄ (Table 2 and Fig. 4b). This could be an important marker for evaluating platelet heterogeneity in sensitive species such as cats. Correlation Matrix Analysis Correlation matrix illustrating the bivariate correlation coefficients (r) between PLT, PCT, MPV, and PDW measured in canine and feline blood using two anticoagulants: K2-EDTA and MgSO4 (Fig. 5a and Fig. 5b). The most striking results involve the analytical consistency between the two anticoagulants (EDTA-'E' and MgSO4-' S'): the correlation between MPV-E and MPV-S was perfect ( r = 1.0). The agreement was also nearly flawless, showing excellent reliability for the PLT (PLT-E and PLT-S at r = 0.97), the PCT (PCT-E and PCT-S at r = 0.92), and the PDW (PDW-E and PDW-S also at r = 0.92). Moving to the physiological relationships, the link between PLT and total mass was powerful, especially within the MgSO4 samples (PLT-S and PCT-S at r = 0.92), and remained strong in the K2-EDTA samples (PLT-E and PCT-E at r = 0.88). Furthermore, the parameters related to platelet size showed a robust association: the correlation between MPV and PDW was uniformly strong for both methods (MPV-E and PDW-E at r = 0.85; MPV-S and PDW-S at r = 0.85). Finally, the relationship between the PLT and the MPV was notably the weakest, registering as moderate: PLT-E and MPV-E correlated at r=0.44, while PLT-S and MPV-S showed a slightly lower r = 0.39 (Fig. 5a). Analysis of the correlation matrix for feline platelet indices reveals a complex structure, including both weak and negative correlations. The strongest positive relationships, as expected, exist between the PLT and PCT. This link was near-perfect for the K2-EDTA sample (PLT-E and PCT-E at r = 0.99) and remained very strong for the MgSO4 sample (PLT-S and PCT-S at r = 0.94). However, the analytical consistency between the two anticoagulants (E vs. S) was variable: PLT showed strong positive agreement ( r = 0.75), while PCT showed moderate to strong agreement ( r = 0.63). In contrast, the MPV showed inferior agreement, with a weak negative correlation (MPV-E and MPV-S at r = -0.11), and the PDW showed very weak agreement ( r = 0.21). Looking at the size metrics, the correlation between PDW-E and MPV-E was moderate and negative ( r = -0.35). Similarly, relationships between PLT and MPV were moderate (PLT-E and MPV-E at r = 0.43) or very weak (PLT-E and MPV-S at r = 0.15). Finally, the relationship between PCT-E and PDW-E was practically zero ( r = −0.048; Fig 5b). Discussion Historically, MgSO4 was recognized as a highly suitable anticoagulant for blood analysis, but it later fell out of favor for routine use following the widespread adoption of EDTA [ 1 , 13 ]. Today, due to renewed scientific interest, MgSO4 is being successfully reestablished as a reliable alternative, particularly for resolving EDTA-induced PTCP and improving the accuracy of automated PLT [ 5 , 9 , 15 ]. EDTA anticoagulant is routinely used for hematological evaluations in veterinary medicine [ 2 ]. However, because of challenges with EDTA in sensitive species, the primary objective of this study was to introduce MgSO4 as a potential anticoagulant for the first time and to evaluate its efficacy for assessing PLT, MPV, PCT, and PDW in dogs and cats. Our results demonstrated species-specific critical differences, with significant implications for laboratory protocols. Our primary finding was a significant increase in mean PLT in feline blood samples collected with MgSO4 (about a 12% increase) compared to those with K2-EDTA. This result has direct clinical relevance in feline hematology. Because of their unique cellular characteristics, feline platelets are prone to severe aggregation in the presence of EDTA [ 17 ]. This aggregation leads to systematic errors in automated counters and to the misdiagnosis of PTCP, thereby compromising diagnostic accuracy [ 16 ]. The observed increase in PLT with MgSO4 use indicates that MgSO4 reduces or inhibits platelet aggregation, thereby providing more accurate and reliable PLT measurements. Human studies show that MgSO4 can serve as an alternative for estimating platelet counts in cases of EDTA-induced [ 3 , 16 ]. Stability assessments have also shown that tube agitation has an acceptable impact on PLT results when using MgSO4, enhancing its potential for routine use [ 4 ]. Mechanistically, the action of EDTA is the chelation of divalent ions, while MgSO4 provides Mg 2+ ions. Subsequent studies in dogs also found that the addition of magnesium chloride (MgCl 2 ) can overcome the effects of EDTA, which interferes with hormone measurements [ 14 ]. Indeed, Mg 2+ ions play a crucial role in protecting blood components from EDTA's chelating effects. This aspect has also been considered in studies combining EDTA and MgSO4 [ 16 ]. The MPV parameter in dogs demonstrated high stability with both anticoagulants (very similar means), confirming MgSO4's ability to preserve platelet morphology and volume in this species, comparable to EDTA [ 9 ]. PDW also yielded nearly identical results in dogs, indicating the high reliability of MgSO4 in measuring platelet size distribution. In cats, although MPV showed a slight difference, PDW with MgSO4 exhibited an 8% increase in the mean and a broader range. This increase in dispersion better reflects the true heterogeneity of platelets in cats, which might be overlooked with EDTA due to platelet aggregation or swelling phenomena [ 9 ]. The importance of PDW as a marker for assessing platelet production dynamics in the bone marrow is well established, and the ability of MgSO4 to capture a broader size spectrum could make it a critical diagnostic marker in sensitive felids, a species in which platelet measurement has long been challenging [ 19 ]. This change in PCT, which is a value calculated from PLT and MPV, provides an indicator for the mass of the platelets, which is particularly relevant for feline species. However, there was a mean increase in PCT of up to 17% with MgSO4 administration. This increase is likely attributable to the elevated PLT and indicates enhanced diagnostic utility, as PCT offers a more accurate assessment of circulating platelet mass in cats. In dogs, a profound decrease in PCT was observed with MgSO4 compared with EDTA. Such a difference may be due to a complex anticoagulant effect, in which the correlation between platelet volume and concentration is involved. Future studies must evaluate MgSO4 alongside other therapeutic variables, such as its analgesic effect in dogs, to provide a well-rounded perspective on its biological effects [ 6 ]. The correlation analysis successfully validated the essential relationship between PLT and PCT (total platelet mass) in feline blood. This link was nearly perfect in K2-EDTA samples ( r = 0.99) and remained exceptionally strong in MgSO4 samples ( r = 0.94). The use of MgSO4 as an alternative anticoagulant is warranted because it is effective in preventing spontaneous in vitro platelet agglutination in anticoagulant-induced PTCP [ 10 ]. The correlation is significant and provides a reliable basis for assessing the reliability of other platelet size measures (MPV and PDW). The results are consistent with other veterinary studies aimed at improving the precision of platelet measurement in cats [ 12 ]. While PLT showed a strong positive agreement ( r = 0.75), PCT showed moderate-to-strong agreement ( r = 0.63). The result indicates that although the PLT can be determined consistently regardless of the anticoagulant used, the PCT calculated is less consistent, which suggests that PCT is more sensitive to the differential effects of anticoagulants, a long-recognized problem of measurement disparity in hematology [ 7 , 8 , 18 ]. Additionally, the weak negative correlation between MPV in K2-EDTA and MgSO4 samples ( r = -0.11) suggests that these anticoagulants do not yield consistent MPV measurements. It is in line with earlier findings where platelet aggregation, common in feline blood, interfered with MPV readings [ 7 ]. Moreover, the moderate negative correlation ( r = -0.35) between PDW and MPV in K2-EDTA samples suggests a compensatory relationship, which could explain variability in platelet size and distribution indices in cats [ 8 ]. The very weak correlation between PCT and PDW ( r = -0.048) in K2-EDTA samples indicates that these two indices are mainly independent. It further supports the complexity of interpreting platelet indices in feline blood, as similar findings have been reported in other studies exploring platelet aggregation and the effects of different anticoagulants [ 8 , 12 ]. In conclusion, MgSO4 is effective as a replacement for K2-EDTA in dogs. MgSO4 in cats is critical because it helps prevent platelet clumping (PTCP), thereby maintaining normal PLT and PCT levels. The beneficial compound in feline blood analysis requires caution in use. There are no favorable MgSO4 values for K2-EDTA with respect to the size parameters MPV and PDW. It suggests such parameters are not consistently accurate, irrespective of the anticoagulant used. Taken together, our findings reveal that MgSO4 may be a reliable in vitro anticoagulant for blood cell counting, as even high agonist concentrations do not induce platelet aggregation in the presence of MgSO4.​ Declarations Acknowledgements This paper is based on the theses of the first and second authors, and the corresponding author would like to express sincere gratitude to Semnan University and Dr. Fattahian's Private Veterinary Hospital for their support and the resources provided throughout this research. Authors’ contributions All authors were involved in the conception and design of the study. M.A-h took the lead in designing and overseeing the research. A.H.T.S, H.G, and M.A-h prepared the materials. M.A-h and H.A carried out data collection and analysis. M.A-h wrote the first draft of the manuscript. All authors provided feedback on earlier drafts of the manuscript and read and approved the final version. Funding No external funding was used for this study. Data availability Raw data is available from the corresponding author upon request. Ethics approval and consent to participate This study received approval from the Semnan University Ethics Committee (IR.SU.REC.1403.24). The relevant ethical committee approved all clinical, experimental, and laboratory protocols. Methodological procedures were strictly complied with national and institutional guidelines and ethical standards. Informed consent was obtained from all pet owners prior to sample collection. Owners were fully informed about the purpose of the study, sampling procedures, potential risks, and benefits, and their right to withdraw at any time. Consent for publication Not applicable. Competing interests The authors declare no competing interests. References Bizzozero J. 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Tables Table 1 Platelet Parameters (PLT, MPV, PCT, PDW) in Canine Blood Samples Anticoagulated with K2-EDTA and MgSO4, Featuring Minimum, Maximum, Mean, Median, and Quartile (25% and 75%) Values Statistic PLT (× 10 3 /µL) MPV (fL) PCT (%) PDW (%) EDTA MgSO 4 EDTA MgSO 4 EDTA MgSO 4 EDTA MgSO 4 min 139.00 53.00 8.00 7.60 0.12 0.05 10.70 8.90 25% 295.00 183.00 9.10 8.70 0.28 0.16 13.30 12.50 Median 369.00 445.00 9.70 9.30 0.37 0.42 14.40 15.70 75% 643.00 952.00 10.40 10.5 0.66 0.93 15.20 17.40 max 1232.0 1250.0 12.00 12.20 1.27 1.31 17.40 21.60 Mean 485.16 545.52 9.78 9.50 0.48 0.56 14.39 15.49 Table 2 Platelet Parameters (PLT, MPV, PCT, PDW) in Feline Blood Samples Anticoagulated with K2-EDTA and MgSO4, Featuring Minimum, Maximum, Mean, Median, and Quartile (25% and 75%) Values Statistic PLT (× 10 3 /µL) MPV (fL) PCT (%) PDW (%) EDTA MgSO 4 EDTA MgSO 4 EDTA MgSO 4 EDTA MgSO 4 min 177.00 183.00 5.60 5.80 0.14 0.11 14.10 14.10 25% 270.25 296.50 6.85 6.70 0.23 0.23 15.12 15.07 Median 394.0 386.0 7.30 7.40 0.28 0.27 15.70 15.65 75% 464.75 476.00 7.85 7.95 0.34 0.35 16.70 16.52 max 956.0 967.0 15.10 15.00 0.73 0.72 17.30 17.30 Mean 404.43 418.63 7.69 7.64 0.56 0.30 15.81 15.78 Additional Declarations No competing interests reported. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8282197","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":597208518,"identity":"7be331f3-6ae6-4a23-bb35-7ce5b70c003c","order_by":0,"name":"Amir Hossein Tofighi Soleimandarabi","email":"","orcid":"","institution":"Semnan University","correspondingAuthor":false,"prefix":"","firstName":"Amir","middleName":"Hossein Tofighi","lastName":"Soleimandarabi","suffix":""},{"id":597208519,"identity":"83de5231-c8be-438c-9318-acff7e2e003b","order_by":1,"name":"Helia Ghasemzadeh","email":"","orcid":"","institution":"Semnan University","correspondingAuthor":false,"prefix":"","firstName":"Helia","middleName":"","lastName":"Ghasemzadeh","suffix":""},{"id":597208520,"identity":"4c5f8d70-9353-4ba1-8333-22f1f53cbdfa","order_by":2,"name":"Mahmood Ahmadi-hamedani","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABG0lEQVRIiWNgGAWjYBACNgkgwWPAkADiANkSckiSjA3YtPCjazEmqEVyBkgLA1wLQyJWZcjA4HZ34oc3BQx5/NMOH7zxc4dF+vz27rQHP3cwyPM3MLd9wKblztnNknMMGIolbqclW/aekcjdcObsdsPeMwyGMw4wNs/ApuVG7gZpoF8SG27nmEnwtgG1SORuAzIYGDcwMDZjc5j9jdzNv0Fa5t/O/yb5t00iXX7+221ABoM9Li1AW7aBbdlwO4dNGmhLAsMN3m1ABlAEjxbLOQYSiRtvpxlby7ZJGG44AzQEyEiecRinls033vyxSZx3O/nhzbdtdfLy7We3Sb5ts7Htb29/jCugGSAxgiHCjEfDKBgFo2AUjAK8AACsSWWoTbRCUgAAAABJRU5ErkJggg==","orcid":"","institution":"Semnan University","correspondingAuthor":true,"prefix":"","firstName":"Mahmood","middleName":"","lastName":"Ahmadi-hamedani","suffix":""},{"id":597208521,"identity":"d496901a-ce92-4866-b132-2e6801996fa8","order_by":3,"name":"Hasti Azarabad","email":"","orcid":"","institution":"Fattahian Private Veterinary Hospital","correspondingAuthor":false,"prefix":"","firstName":"Hasti","middleName":"","lastName":"Azarabad","suffix":""}],"badges":[],"createdAt":"2025-12-04 19:23:17","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8282197/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8282197/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":103536780,"identity":"78b36694-d096-4ebc-9e48-763ffecb44fb","added_by":"auto","created_at":"2026-02-26 18:42:52","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":187070,"visible":true,"origin":"","legend":"\u003cp\u003eBox Plot comparing the total platelet counts (PLT, ×10\u003csup\u003e3\u003c/sup\u003e/μL on the Y-axis) measured in canine (a) and feline (b) blood samples collected in K2-EDTA (PLT-E) and MgSO4\u003cbr\u003e\n(PLT-S) tubes. The boxes represent the interquartile range (IQR), the central horizontal line indicates the median, and the whiskers denote the maximum and minimum values excluding outliers (individual points/diamonds).\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8282197/v1/10f706b8091d58645f5fc2ca.jpeg"},{"id":103536784,"identity":"64fa57c5-16e5-4c7c-8ae2-6b2ad18cdef5","added_by":"auto","created_at":"2026-02-26 18:42:52","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":333991,"visible":true,"origin":"","legend":"\u003cp\u003eBox Plot comparing the mean platelet volume (MPV, fL on the Y-axis) measured in canine (a) and feline (b) blood samples collected in K2-EDTA (MPV-E) and MgSO4 (MPV-S) tubes. The boxes represent the interquartile range (IQR), the central horizontal line indicates the median, and the whiskers denote the maximum and minimum values excluding outliers (individual points/diamonds).\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8282197/v1/a6e0254e52f85f45a2e6bd24.jpeg"},{"id":104398818,"identity":"08305d80-0147-4918-b4e8-42614326e4b7","added_by":"auto","created_at":"2026-03-11 12:03:44","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":89155,"visible":true,"origin":"","legend":"\u003cp\u003eBox Plot comparing the plateletcrit (PCT, % on the Y-axis) measured in canine (a) and feline (b) blood samples collected in K2-EDTA (PCT-E) and MgSO4 (PCT-S) tubes. The boxes represent the interquartile range (IQR), the central horizontal line indicates the median, and the whiskers denote the maximum and minimum values excluding outliers (individual points/diamonds).\u003c/p\u003e","description":"","filename":"floatimage3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8282197/v1/2a03963a18ce0afb23854999.jpeg"},{"id":104398385,"identity":"063cc78e-06bb-4164-93a6-f5e2932affcb","added_by":"auto","created_at":"2026-03-11 12:02:06","extension":"jpeg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":106617,"visible":true,"origin":"","legend":"\u003cp\u003eBox Plot comparing the platelet distribution width (PDW, % on the Y-axis) measured in canine (a) and feline (b) blood samples collected in K2-EDTA (PDW-E) and MgSO4 (PDW-S) tubes. The boxes represent the interquartile range (IQR), the central horizontal line indicates the median, and the whiskers denote the maximum and minimum values excluding outliers (individual points/diamonds).\u003c/p\u003e","description":"","filename":"floatimage4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8282197/v1/274ec3ce4c2c8876b84e107c.jpeg"},{"id":103536783,"identity":"1560c17c-0a39-4c57-a41d-6fadc5d7bbe2","added_by":"auto","created_at":"2026-02-26 18:42:52","extension":"jpeg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":249571,"visible":true,"origin":"","legend":"\u003cp\u003eCorrelation matrix illustrating the bivariate correlation coefficients (r) between platelet count (PLT), Plateletcrit (PCT), Mean Platelet Volume (MPV), and Platelet Distribution Width (PDW) measured in canine (a) and feline (b) blood using two anticoagulants: K2-EDTA (indicated by the suffix 'E') and MgSO4​ (indicated by the suffix 'S'). For straightforward interpretation, the size of each circle directly reflects the absolute strength of the correlation (abs (value)). At the same time, the color intensity (from light pink to bright red) clearly shows the sign and magnitude of that relationship (value), meaning a larger, darker red circle represents a stronger positive connection.\u003c/p\u003e","description":"","filename":"floatimage5.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8282197/v1/302ef7a4e2f3f990323a8420.jpeg"},{"id":109093270,"identity":"3b5a080f-9c71-48d5-9c52-2c1f42c5aab0","added_by":"auto","created_at":"2026-05-12 13:46:13","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1276253,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8282197/v1/1282084b-9a46-43f3-8910-2c41da61ba0e.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Assessing the analytical reliability of platelet counts and related indices in dogs and cats: a correlation matrix analysis comparing K2-EDTA and MgSO4 anticoagulation","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAccurately measuring platelet parameters, such as platelet count (PLT), mean platelet volume (MPV), plateletcrit (PCT), and platelet distribution width (PDW), is essential for evaluating coagulation, inflammation, and thrombotic risk in veterinary medicine. It is commonly accepted that in hematology laboratories, ethylene diamine tetra-acetic acid (EDTA) becomes the preferred anticoagulant due to its ability to chelate calcium and, consequently, inhibit coagulation development [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. However, the use of EDTA can also lead to pre-analytical issues that invalidate test results. In humans, a rare side effect of EDTA is pseudothrombocytopenia (PTCP), in which the platelet count appears low because platelets clump together in the test tube. It occurs in about 0.07\u0026ndash;0.27% of EDTA samples and is most often observed when samples are kept at room temperature [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Platelet aggregation is also a characteristic feature of canine and feline blood samples that can lead to misleading diagnoses of thrombocytopenia and to the inappropriate completion of diagnostic profiles or the supposed treatment process [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Cold agglutinin antibodies, autoantibody precipitation upon addition of EDTA, or low anticoagulant levels or platelet activation can result in platelet aggregation in a venous blood sample [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. The development and progression of this process can differ among different species. It is less observed in dogs than in cats. Aggregation by platelets would be more common in felines, and the process could be found out easily due to the unique properties of feline platelets, such as their osmotic influence of EDTA and relatively smaller size [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. This clumping may lead to inaccurate platelet counts, slowing the diagnostic test for thrombocytopenia. EDTA may also alter the cell's osmotic pressure, causing swelling that can make platelet volume measurements, such as MPV, appear higher than they really are [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Some studies in humans have shown that EDTA-PTCP is associated with IgG autoantibodies targeting a hidden region of the IIb unit of the GPIIbIIIa receptor (also known as CD41/CD61), which is revealed only in the presence of EDTA [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Researchers use three main ways to reduce the effects of PTCP in dogs and cats. The first is to prewarm refrigerated samples before analysis. The second is to add PLT antagonists, such as prostaglandin E1 (PGE1), citrate, theophylline, adenosine, dipyridamole (CTAD), and Iloprost, to collection tubes before drawing blood [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. The third intervention involves pretreating EDTA tubes with amikacin, which neutralizes the fibrinogen receptor GPIIbIIIa and reduces its fibrinogen binding [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. To reduce these pre-analytical problems, especially in preparations that are likely to exhibit platelet aggregation, other anticoagulants, including magnesium sulfate (MgSO4), are also added. MgSO4 works as an anticoagulant because magnesium ions compete with calcium in coagulation reactions and help inhibit clotting [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. More importantly, MgSO4 is also recognized as an antiplatelet aggregation agent; magnesium, being a calcium antagonist, can inhibit platelet activation and prevent clumping [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Studies in people have shown that MgSO4 improves PLT function in those with PTCP. It means MgSO4 can give more true results [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Right now, there is no other choice like MgSO4 for EDTA-PTCP that can deliver better or more accurate results for dogs and cats. Given the species-specific challenges and the use of the specific hematology analyzer, Celtac-Alpha (Nihon Kohden), in our study, the main objective of this research is to compare the discrepancies and statistical correlations of PLT and related platelet indices between K2-EDTA and MgSO4 anticoagulants in dog and cat blood samples.\u003c/p\u003e"},{"header":"Material and methods","content":"\u003cp\u003e\u003cstrong\u003eStudied animals and sample collection\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe conducted a cross-sectional study utilizing dogs and cats that presented for routine health check-ups at a private veterinary hospital in Tehran, Iran. A total of 61 client-owned dogs and cats (30 dogs and 31 cats) were ultimately selected for this study. All animals included in the study were clinically healthy, with no history of coagulation disorders, underlying platelet diseases, or use of anticoagulants or nonsteroidal anti-inflammatory drugs (NSAIDs). For each dog and cat included in the study, essential demographic data such as age, sex, breed, and the individual\u0026apos;s name were carefully recorded along with the blood sample. No animals were euthanized or sacrificed for this study. Blood samples were collected from live animals as non-invasive clinical specimens, with no anesthesia required; the animals remained fully conscious throughout the collection process.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAnticoagulants used\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFor this study, we collected paired, simultaneous blood samples from 31 cats and 30 dogs (a total of 122 tubes) immediately after venipuncture. We used two types of tubes: K2-EDTA (S-Monovette K2-EDTA; fill volume 2.6 mL) as the routine standard for hematology, and MgSO4 (S-Monovette ThromboExact; fill volume 2.7 mL) [10]. The MgSO4 collection tubes were particularly selected as an anticoagulant method alternative based on research work that proposes it assists in combating PTCP, which stands for the phenomenon wherein the clotting agent EDTA causes the platelet cell masses to aggregate, thus producing an inflated cell count [3, 16].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePre-analytical phase\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo prevent platelet preactivation, the blood was fully extended after the tourniquet was released, gently mixed with the anticoagulant, and immediately brought to the laboratory for further processing [10]. Platelet-related measurements, particularly sample storage, may result in volume changes in K2-EDTA. Aggregation, a property seen with feline platelets, would be enhanced [19]. To prevent unwanted changes, all hematological analyses were performed within 30 minutes of blood collection [9].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMeasurement of platelet parameters\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePLT and platelet indices, including MPV, PCT, and PDW, were measured using the Celltac \u0026alpha; VET MEK-6550 hematology analyzer (Nihon Kohden). This device operates using the impedance method and can accurately measure these parameters in canine and feline blood [9]. PCT was calculated as a function of PLT and MPV.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMeasurements for each parameter were recorded in paired sets using both the K2-EDTA and MgSO4 anticoagulants to enable direct comparison. Initially, the Shapiro-Wilk test was performed on each parameter to assess its data distribution and normality. Since the majority of parameters (PLT, MPV, and PCT) did not exhibit a normal distribution in either dog or cat samples, the Wilcoxon Signed-Rank Test, a non-parametric method, was strategically employed to compare the effects of K2-EDTA versus MgSO4 on these values. Conversely, because the PDW parameter did follow a normal distribution, its mean values were compared using the Paired Samples t-Test, a parametric test. To evaluate the level of agreement between the two anticoagulants and to determine the relationships among all measured parameters, a Pearson\u0026rsquo;s Correlation Coefficient matrix was calculated. A strong correlation, close to 1, was considered representative of strong agreement between variables, implying the possibility of interchangeability. Data analysis and graphics processing were carried out in R-Studio (version 4.4.2). A p-value less than 0.05 was considered statistically significant.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eThe platelet parameters, such as PLT, MPV, PCT, and PDW, were evaluated in whole-blood samples from 30 dogs and 31 cats using two anticoagulants: K2-EDTA (E) and MgSO4 (S). The exact statistical distribution, minimum, Q1, median, Q3, maximum, and mean, is given for each parameter in Tables 1 and 2.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eComparison of PLT between K2-EDTA and MgSO4 anticoagulants in dogs and cats\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn the PLT range in K2-EDTA, from 177 to 956, the mean was 404.43, with a median of 394.0; in MgSO4, the range was similar, from 183 to 967, but the mean was slightly higher, at 418.63, with a median of 386.0. Such an increase in mean could indicate reduced platelet aggregation in MgSO4 (Table 1 and Fig. 1a). This parameter varied more in cats: in K2-EDTA, it ranged from 139 to 1232 (mean 485.16, median 369.00), whereas in MgSO4, the mean increased to 545.52 (range 53 to 1250, median 445.00). This difference of about 12% may be important for improving diagnostic accuracy in cats, where PLT is generally variable due to aggregation (Table 2 and Fig. 1b).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eComparison of MPV between K2-EDTA and MgSO4 anticoagulants in dogs and cats\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn dogs, MPV was found to be highly stable, with a range of 5.60\u0026ndash;15.10 in K2-EDTA (mean: 7.69, median: 7.30) and a comparably stable range in MgSO4 (mean: 7.64, median: 7.40), thus highlighting the MgSO4 anticoagulant\u0026rsquo;s strong capacity for maintaining platelet morphology (Table 1, Fig. 2a). In the case of cats, MPV was higher, with a range of 8.00\u0026ndash;12.00 in K2-EDTA (mean: 9.78, median: 9.70) and 7.60 to 12.20 in MgSO4 (mean 9.50, median 9.30). This slight difference could indicate species-specific effects of anticoagulants on platelet volume, underscoring the importance of adjusting laboratory protocols (Table 2 and Fig. 2b).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eComparison of PCT between K2-EDTA and MgSO4 anticoagulants in dogs and cats\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn dogs, PCT ranged from 0.14 to 0.73 in K2-EDTA with a mean of 0.56 and a median of 0.28, while in MgSO4, the mean decreased to 0.30 (range 0.11 to 0.72, median 0.27). This significant difference might reflect differences in the overall effect on platelet volume (Table 1 and Fig. 3a). In cats, this parameter increased significantly more in MgSO4; in K2-EDTA, the range was 0.12 to 1.27, with an average of 0.48 and a median of 0.37, while in MgSO4, the values ranged from 0.05 to 1.31, with an average value of 0.56 and a median of 0.42. An increase of 17% may represent one of the main discoveries for improved diagnostic performance in cats, underscoring the potential of MgSO4 to reduce variability (Table 2 and Fig. 3b).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eComparison of PDW between K2-EDTA and MgSO4 anticoagulants in dogs and cats\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePDW in dogs was very similar, with a range of 14.10 to 17.30 in both anticoagulants (mean approximately 15.8 in both; median around 15.7), strongly emphasizing the reliability of MgSO₄ in measuring platelet size distribution (Table 1 and Fig. 4a). In cats, there was a larger spread: the range in EDTA was from 10.70 to 17.40 (mean 14.39, median 14.40), while in MgSO₄, it ranged from 8.90 to 21.60 (mean 15.49, median 15.70). The approximately 8% increase in mean platelet size suggests greater variation in the platelet size distribution under MgSO₄ (Table 2 and Fig. 4b). This could be an important marker for evaluating platelet heterogeneity in sensitive species such as cats.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCorrelation Matrix Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCorrelation matrix illustrating the bivariate correlation coefficients (r) between PLT, PCT, MPV, and PDW measured in canine and feline blood using two anticoagulants: K2-EDTA and MgSO4\u003cspan dir=\"RTL\"\u003e\u0026nbsp;\u003c/span\u003e(Fig. 5a and Fig. 5b).\u0026nbsp;The most striking results involve the analytical consistency between the two anticoagulants (EDTA-\u0026apos;E\u0026apos; and MgSO4-\u0026apos; S\u0026apos;): the correlation between MPV-E and MPV-S was perfect (\u003cem\u003er\u003c/em\u003e = 1.0). The agreement was also nearly flawless, showing excellent reliability for the PLT (PLT-E and PLT-S at \u003cem\u003er\u003c/em\u003e = 0.97), the PCT (PCT-E and PCT-S at \u003cem\u003er\u003c/em\u003e = 0.92), and the PDW (PDW-E and PDW-S also at \u003cem\u003er\u003c/em\u003e = 0.92). Moving to the physiological relationships, the link between PLT and total mass was powerful, especially within the MgSO4 samples (PLT-S and PCT-S at \u003cem\u003er\u003c/em\u003e = 0.92), and remained strong in the K2-EDTA samples (PLT-E and PCT-E at \u003cem\u003er\u0026nbsp;\u003c/em\u003e= 0.88). Furthermore, the parameters related to platelet size showed a robust association: the correlation between MPV and PDW was uniformly strong for both methods (MPV-E and PDW-E at \u003cem\u003er\u003c/em\u003e = 0.85; MPV-S and PDW-S at \u003cem\u003er\u003c/em\u003e = 0.85). Finally, the relationship between the PLT and the MPV was notably the weakest, registering as moderate: PLT-E and MPV-E correlated at r=0.44, while PLT-S and MPV-S showed a slightly lower \u003cem\u003er\u003c/em\u003e = 0.39 (Fig. 5a).\u003c/p\u003e\n\u003cp\u003eAnalysis of the correlation matrix for feline platelet indices reveals a complex structure, including both weak and negative correlations. The strongest positive relationships, as expected, exist between the PLT and PCT. This link was near-perfect for the K2-EDTA sample (PLT-E and PCT-E at \u003cem\u003er\u003c/em\u003e = 0.99) and remained very strong for the MgSO4 sample (PLT-S and PCT-S at \u003cem\u003er\u003c/em\u003e = 0.94). However, the analytical consistency between the two anticoagulants (E vs. S) was variable: PLT showed strong positive agreement (\u003cem\u003er\u003c/em\u003e = 0.75), while PCT showed moderate to strong agreement (\u003cem\u003er\u003c/em\u003e = 0.63). In contrast, the MPV showed inferior agreement, with a weak negative correlation (MPV-E and MPV-S at \u003cem\u003er\u003c/em\u003e = -0.11), and the PDW showed very weak agreement (\u003cem\u003er\u003c/em\u003e = 0.21). Looking at the size metrics, the correlation between PDW-E and MPV-E was moderate and negative (\u003cem\u003er\u0026nbsp;\u003c/em\u003e= -0.35). Similarly, relationships between PLT and MPV were moderate (PLT-E and MPV-E at \u003cem\u003er\u003c/em\u003e = 0.43) or very weak (PLT-E and MPV-S at \u003cem\u003er\u003c/em\u003e = 0.15). Finally, the relationship between PCT-E and PDW-E was practically zero (\u003cem\u003er\u003c/em\u003e = \u0026minus;0.048; Fig 5b).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eHistorically, MgSO4 was recognized as a highly suitable anticoagulant for blood analysis, but it later fell out of favor for routine use following the widespread adoption of EDTA [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. Today, due to renewed scientific interest, MgSO4 is being successfully reestablished as a reliable alternative, particularly for resolving EDTA-induced PTCP and improving the accuracy of automated PLT [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. EDTA anticoagulant is routinely used for hematological evaluations in veterinary medicine [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. However, because of challenges with EDTA in sensitive species, the primary objective of this study was to introduce MgSO4 as a potential anticoagulant for the first time and to evaluate its efficacy for assessing PLT, MPV, PCT, and PDW in dogs and cats. Our results demonstrated species-specific critical differences, with significant implications for laboratory protocols. Our primary finding was a significant increase in mean PLT in feline blood samples collected with MgSO4 (about a 12% increase) compared to those with K2-EDTA. This result has direct clinical relevance in feline hematology. Because of their unique cellular characteristics, feline platelets are prone to severe aggregation in the presence of EDTA [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. This aggregation leads to systematic errors in automated counters and to the misdiagnosis of PTCP, thereby compromising diagnostic accuracy [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. The observed increase in PLT with MgSO4 use indicates that MgSO4 reduces or inhibits platelet aggregation, thereby providing more accurate and reliable PLT measurements. Human studies show that MgSO4 can serve as an alternative for estimating platelet counts in cases of EDTA-induced [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Stability assessments have also shown that tube agitation has an acceptable impact on PLT results when using MgSO4, enhancing its potential for routine use [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Mechanistically, the action of EDTA is the chelation of divalent ions, while MgSO4 provides Mg\u003csup\u003e2+\u003c/sup\u003e ions. Subsequent studies in dogs also found that the addition of magnesium chloride (MgCl\u003csub\u003e2\u003c/sub\u003e) can overcome the effects of EDTA, which interferes with hormone measurements [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Indeed, Mg\u003csup\u003e2+\u003c/sup\u003e ions play a crucial role in protecting blood components from EDTA's chelating effects. This aspect has also been considered in studies combining EDTA and MgSO4 [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. The MPV parameter in dogs demonstrated high stability with both anticoagulants (very similar means), confirming MgSO4's ability to preserve platelet morphology and volume in this species, comparable to EDTA [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. PDW also yielded nearly identical results in dogs, indicating the high reliability of MgSO4 in measuring platelet size distribution. In cats, although MPV showed a slight difference, PDW with MgSO4 exhibited an 8% increase in the mean and a broader range. This increase in dispersion better reflects the true heterogeneity of platelets in cats, which might be overlooked with EDTA due to platelet aggregation or swelling phenomena [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. The importance of PDW as a marker for assessing platelet production dynamics in the bone marrow is well established, and the ability of MgSO4 to capture a broader size spectrum could make it a critical diagnostic marker in sensitive felids, a species in which platelet measurement has long been challenging [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. This change in PCT, which is a value calculated from PLT and MPV, provides an indicator for the mass of the platelets, which is particularly relevant for feline species. However, there was a mean increase in PCT of up to 17% with MgSO4 administration. This increase is likely attributable to the elevated PLT and indicates enhanced diagnostic utility, as PCT offers a more accurate assessment of circulating platelet mass in cats. In dogs, a profound decrease in PCT was observed with MgSO4 compared with EDTA. Such a difference may be due to a complex anticoagulant effect, in which the correlation between platelet volume and concentration is involved. Future studies must evaluate MgSO4 alongside other therapeutic variables, such as its analgesic effect in dogs, to provide a well-rounded perspective on its biological effects [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe correlation analysis successfully validated the essential relationship between PLT and PCT (total platelet mass) in feline blood. This link was nearly perfect in K2-EDTA samples (\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.99) and remained exceptionally strong in MgSO4 samples (\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.94). The use of MgSO4 as an alternative anticoagulant is warranted because it is effective in preventing spontaneous in vitro platelet agglutination in anticoagulant-induced PTCP [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. The correlation is significant and provides a reliable basis for assessing the reliability of other platelet size measures (MPV and PDW). The results are consistent with other veterinary studies aimed at improving the precision of platelet measurement in cats [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. While PLT showed a strong positive agreement (\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.75), PCT showed moderate-to-strong agreement (\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.63). The result indicates that although the PLT can be determined consistently regardless of the anticoagulant used, the PCT calculated is less consistent, which suggests that PCT is more sensitive to the differential effects of anticoagulants, a long-recognized problem of measurement disparity in hematology [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. Additionally, the weak negative correlation between MPV in K2-EDTA and MgSO4 samples (\u003cem\u003er\u003c/em\u003e = -0.11) suggests that these anticoagulants do not yield consistent MPV measurements. It is in line with earlier findings where platelet aggregation, common in feline blood, interfered with MPV readings [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Moreover, the moderate negative correlation (\u003cem\u003er\u003c/em\u003e = -0.35) between PDW and MPV in K2-EDTA samples suggests a compensatory relationship, which could explain variability in platelet size and distribution indices in cats [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. The very weak correlation between PCT and PDW (\u003cem\u003er\u003c/em\u003e = -0.048) in K2-EDTA samples indicates that these two indices are mainly independent. It further supports the complexity of interpreting platelet indices in feline blood, as similar findings have been reported in other studies exploring platelet aggregation and the effects of different anticoagulants [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn conclusion, MgSO4 is effective as a replacement for K2-EDTA in dogs. MgSO4 in cats is critical because it helps prevent platelet clumping (PTCP), thereby maintaining normal PLT and PCT levels. The beneficial compound in feline blood analysis requires caution in use. There are no favorable MgSO4 values for K2-EDTA with respect to the size parameters MPV and PDW. It suggests such parameters are not consistently accurate, irrespective of the anticoagulant used. Taken together, our findings reveal that MgSO4 may be a reliable in vitro anticoagulant for blood cell counting, as even high agonist concentrations do not induce platelet aggregation in the presence of MgSO4.​\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis paper is based on the theses of the first and second authors, and the corresponding author would like to express sincere gratitude to Semnan University and Dr. Fattahian\u0026apos;s Private Veterinary Hospital for their support and the resources provided throughout this research.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026rsquo; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors were involved in the conception and design of the study. M.A-h took the lead in designing and overseeing the research. A.H.T.S, H.G, and M.A-h prepared the materials. M.A-h and H.A carried out data collection and analysis. M.A-h wrote the first draft of the manuscript. All authors provided feedback on earlier drafts of the manuscript and read and approved the final version.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNo external funding was used for this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eRaw data is available from the corresponding author upon request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study received approval from the Semnan University Ethics Committee (IR.SU.REC.1403.24). The relevant ethical committee approved all clinical, experimental, and laboratory protocols. Methodological procedures were strictly complied with national and institutional guidelines and ethical standards. Informed consent was obtained from all pet owners prior to sample collection. Owners were fully informed about the purpose of the study, sampling procedures, potential risks, and benefits, and their right to withdraw at any time.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e"},{"header":"References","content":"\u003col start=\"1\" type=\"1\"\u003e\n \u003cli\u003eBizzozero J. Ueber einen neuen Formbestandtheil des Blutes und dessen Rolle bei der Thrombose und der Blutgerinnung: Untersuchungen. Arch Pathol Anat Physiol Klin Med. 1882;90(2):261-332.\u003c/li\u003e\n \u003cli\u003eCaudill MN, Meichner K, Koenig A, Berghaus RD, Garner B. Comparison of serum versus EDTA plasma in canine major crossmatch reactions. Vet Clin Pathol. 2021;50(3):319-326.\u003c/li\u003e\n \u003cli\u003eChoccalingam C, Kanna Nandagopal Radha R, Snigdha N. Estimation of Platelet Counts and Other Hematological Parameters in Pseudothrombocytopenia Using Alternative Anticoagulant: Magnesium Sulfate. Clin Med Insights Blood Disord. 2017;10:1179545X17705380.\u003c/li\u003e\n \u003cli\u003eEvans DP, Tariq M, Sujata B, McCann G, Sobki S. The effects of magnesium sulphate and EDTA in the hypercholesterolaemic rabbit. Diabetes Obes Metab. 2001;3(6):417-422.\u003c/li\u003e\n \u003cli\u003eFran\u0026ccedil;ois D, Masure A, Atallah N, Touil L, Vasse M. Underestimation of platelet count on magnesium salt-anticoagulated samples. Clin Chem Lab Med. 2014;52(5):e95-e97.\u003c/li\u003e\n \u003cli\u003eGalosi M, Pennasilico L, Piccionello AP, Serino F, Tosi F, Sassaroli S, Di Bella C. Effects of a synergic interaction between magnesium sulphate and ketamine on the perioperative nociception in dogs undergoing tibial plateau leveling osteotomy: a pilot study. Front Vet Sci. 2024;11:1453673.\u003c/li\u003e\n \u003cli\u003eGranat F, Geffr\u0026eacute; A, Braun JP, Trumel C. Comparison of platelet clumping and complete blood count results with Sysmex XT-2000iV in feline blood sampled on EDTA or EDTA plus CTAD (citrate, theophylline, adenosine and dipyridamole). J Feline Med Surg. 2011;13(12):953-958.\u003c/li\u003e\n \u003cli\u003eHo KK, Abrams-Ogg AC, Wood RD, O\u0026rsquo;Sullivan ML, Kirby GM, Blois SL. Assessment of platelet function in healthy sedated cats using three whole blood platelet function tests. J Vet Diagn Invest. 2015;27(3):352-360.\u003c/li\u003e\n \u003cli\u003eMannu\u0026szlig; S, Schuff-Werner P, Drei\u0026szlig;iger K, Kohlschein P. Magnesium sulfate as an alternative in vitro anticoagulant for the measurement of platelet parameters? Am J Clin Pathol. 2016;145(6):806-814.\u003c/li\u003e\n \u003cli\u003eMannu\u0026szlig; S, Schuff-Werner P, Drei\u0026szlig;iger K, Burstein C. Inhibition of agonist-induced platelet aggregation by magnesium sulfate warrants its use as an alternative in vitro anticoagulant in pseudothrombocytopenia. Platelets. 2020;31(5):680-684.\u003c/li\u003e\n \u003cli\u003eMylonakis ME, Leontides L, Farmaki R, Kostoulas P, Koutinas AF, Christopher M. Effect of anticoagulant and storage conditions on platelet size and clumping in healthy dogs. J Vet Diagn Invest. 2008;20(6):774-779.\u003c/li\u003e\n \u003cli\u003eNorman EJ, Barron RC, Nash AS, Clampitt RB. Prevalence of low automated platelet counts in cats: comparison with prevalence of thrombocytopenia based on blood smear estimation. Vet Clin Pathol. 2001;30(3):137-140.\u003c/li\u003e\n \u003cli\u003eRavn HB, Kristensen SD, Vissinger H, Husted SE. Magnesium inhibits human platelets. Blood Coagul Fibrinolysis. 1996;7(2):241-244.\u003c/li\u003e\n \u003cli\u003eSchechter DA, Lee HP, Kemppainen RJ, Behrend EN. Effect of EDTA on measurement of cortisol and thyroxine by chemiluminescent enzyme immunoassay in dogs. J Vet Diagn Invest. 2020;32(3):363-368.\u003c/li\u003e\n \u003cli\u003eSchuff-Werner P, Steiner M, Fenger S, Gross HJ, Bierlich A, Dreissiger K, Kohlschein P. Effective estimation of correct platelet counts in pseudothrombocytopenia using an alternative anticoagulant based on magnesium salt. Br J Haematol. 2013;162(5):684-692.\u003c/li\u003e\n \u003cli\u003eSoulard M, Croix P, Cohen P. Comparison of platelet count results on the Sysmex XN between citrate or MgSO4 and K2 EDTA anticoagulants. Int J Lab Hematol. 2023;45(1):20-28.\u003c/li\u003e\n \u003cli\u003eSoulard M, Ketatni H, Visseaux C, Croix P, Cohen P. Impact of shaking EDTA, citrate, or MgSO4 tubes on platelet count results. J Clin Med. 2024;13(18):5350.\u003c/li\u003e\n \u003cli\u003eStokol T, Erb HN. A comparison of platelet parameters in EDTA‐and citrate‐anticoagulated blood in dogs. Vet Clin Pathol. 2007;36(2):148-154.\u003c/li\u003e\n \u003cli\u003eTvedten HW, Ljusner J, Lillieh\u0026ouml;\u0026ouml;k IE. Enumeration of feline platelets in ethylenediamine tetra-acetic acid anticoagulated blood with the ADVIA 2120 system and two manual methods: Leucoplate and Thrombo-TIC. J Vet Diagn Invest. 2013;25(4):493-497.\u003c/li\u003e\n \u003cli\u003eVasilatis DM, Walker NJ, Borjesson DL. Amikacin disaggregates platelet clumps in EDTA blood samples from cats and dogs when added postcollection. Vet Clin Pathol. 2023;52(2):228-235.\u003c/li\u003e\n \u003cli\u003eWilliams TL, Archer J. Effect of prewarming EDTA blood samples to 37\u0026deg; C on platelet count measured by Sysmex XT‐2000iV in dogs, cats, and horses. Vet Clin Pathol. 2016;45(3):444-449.\u003c/li\u003e\n \u003cli\u003eZafar H, Shang Y, Li J, David GA III, Fernandez JP, Molina H, Coller BS. \u0026alpha;IIb\u0026beta;3 binding to a fibrinogen fragment lacking the \u0026gamma;-chain dodecapeptide is activation dependent and EDTA inducible. Blood Adv. 2017;1(7):417-428.\u003c/li\u003e\n \u003cli\u003eZanfagnini LG, Kassab S, Campos D, de Oliveira Chaves JK, Pacheco AD. How much does the platelet aggregate influence the total leukocyte count? Comparison between manual and automated impedance methods in domestic cats. Acta Sci Vet. 2021;49:e799.\u003c/li\u003e\n\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003e\u003cstrong\u003eTable 1\u003c/strong\u003e Platelet Parameters (PLT, MPV, PCT, PDW) in Canine Blood Samples Anticoagulated with K2-EDTA and MgSO4, Featuring Minimum, Maximum, Mean, Median, and Quartile (25% and 75%) Values\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" align=\"\" width=\"107%\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 9px;\"\u003e\n \u003cp\u003eStatistic\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 20px;\"\u003e\n \u003cp\u003ePLT (\u0026times; 10\u003csup\u003e3\u003c/sup\u003e/\u0026micro;L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 20px;\"\u003e\n \u003cp\u003eMPV (fL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 20px;\"\u003e\n \u003cp\u003ePCT (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 16px;\"\u003e\n \u003cp\u003ePDW (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"bottom\" style=\"width: 7px;\"\u003e\n \u003cp\u003eEDTA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12px;\"\u003e\n \u003cp\u003eMgSO\u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 7px;\"\u003e\n \u003cp\u003eEDTA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12px;\"\u003e\n \u003cp\u003eMgSO\u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 7px;\"\u003e\n \u003cp\u003eEDTA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 12px;\"\u003e\n \u003cp\u003eMgSO\u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 7px;\"\u003e\n \u003cp\u003eEDTA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 8px;\"\u003e\n \u003cp\u003eMgSO\u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003emin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e139.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e53.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e8.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e7.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e0.12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e10.70\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 8px;\"\u003e\n \u003cp\u003e8.90\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003e25%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e295.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e183.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e9.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e8.70\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e0.28\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e0.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e13.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 8px;\"\u003e\n \u003cp\u003e12.50\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003eMedian\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e369.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e445.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e9.70\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e9.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e0.37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e0.42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e14.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 8px;\"\u003e\n \u003cp\u003e15.70\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003e75%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e643.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e952.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e10.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e10.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e0.66\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e0.93\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e15.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 8px;\"\u003e\n \u003cp\u003e17.40\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003emax\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e1232.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e1250.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e12.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e12.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e1.27\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e1.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e17.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 8px;\"\u003e\n \u003cp\u003e21.60\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003eMean\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e485.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e545.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e9.78\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e9.50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e0.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e0.56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 4px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 7px;\"\u003e\n \u003cp\u003e14.39\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 8px;\"\u003e\n \u003cp\u003e15.49\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2\u003c/strong\u003e Platelet Parameters (PLT, MPV, PCT, PDW) in Feline Blood Samples Anticoagulated with K2-EDTA and MgSO4, Featuring Minimum, Maximum, Mean, Median, and Quartile (25% and 75%) Values\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" align=\"\" width=\"108%\" class=\"fr-table-selection-hover\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 9px;\"\u003e\n \u003cp\u003eStatistic\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"bottom\" style=\"width: 20px;\"\u003e\n \u003cp\u003ePLT (\u0026times; 10\u003csup\u003e3\u003c/sup\u003e/\u0026micro;L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"bottom\" style=\"width: 20px;\"\u003e\n \u003cp\u003eMPV (fL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"bottom\" style=\"width: 20px;\"\u003e\n \u003cp\u003ePCT (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"bottom\" style=\"width: 17px;\"\u003e\n \u003cp\u003ePDW (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003eEDTA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 11px;\"\u003e\n \u003cp\u003eMgSO\u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003eEDTA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 12px;\"\u003e\n \u003cp\u003eMgSO\u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003eEDTA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 12px;\"\u003e\n \u003cp\u003eMgSO\u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003eEDTA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 9px;\"\u003e\n \u003cp\u003eMgSO\u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003emin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e177.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 11px;\"\u003e\n \u003cp\u003e183.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e5.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 12px;\"\u003e\n \u003cp\u003e5.80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e0.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 12px;\"\u003e\n \u003cp\u003e0.11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e14.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 9px;\"\u003e\n \u003cp\u003e14.10\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003e25%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e270.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 11px;\"\u003e\n \u003cp\u003e296.50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e6.85\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 12px;\"\u003e\n \u003cp\u003e6.70\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e0.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 12px;\"\u003e\n \u003cp\u003e0.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e15.12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 9px;\"\u003e\n \u003cp\u003e15.07\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 9px;\"\u003e\n \u003cp\u003eMedian\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e394.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 11px;\"\u003e\n \u003cp\u003e386.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e7.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 12px;\"\u003e\n \u003cp\u003e7.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 3px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 8px;\"\u003e\n \u003cp\u003e0.28\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 12px;\"\u003e\n \u003cp\u003e0.27\u003c/p\u003e\n \u003c/td\u003e\n 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\u003c/tbody\u003e\n\u003c/table\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Platelet Counts, EDTA, MgSO4, Dogs, Cats","lastPublishedDoi":"10.21203/rs.3.rs-8282197/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8282197/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe standard anticoagulant, ethylene diamine tetra-acetic acid (EDTA), often produces inaccurate platelet results (platelet count (PLT), plateletcrit (PCT), mean platelet volume (MPV), and platelet distribution width (PDW) in dog and cat blood due to its tendency to induce aggregation. This research, therefore, proposes magnesium sulfate (MgSO4) as the superior alternative because it actively prevents this clumping. The research examined blood samples from 31 feline and 32 canine individuals, totaling 122 blood tubes. Two types of tubes were used: k2-EDTA and MgSO4. Clinically healthy dogs and cats were used to obtain the samples, and PLT levels and related indices, such as MPV, PCT, and PDW, were measured using the Celltac α VET MEK-6550 hematology analyzer. In dogs, platelet parameters in MgSO4 and EDTA showed strong agreement (\u003cem\u003er\u003c/em\u003e\u0026thinsp;\u0026ge;\u0026thinsp;0.90 for MPV, PLT, PCT, and PDW), with stability in MPV but a decrease in mean PCT in MgSO4 (0.30 vs. 0.56 in K2-EDTA). Feline results confirmed a strong correlation between PLT and PCT (\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.99), but analytical agreement between K2-EDTA and MgSO4 was poor for MPV (\u003cem\u003er\u003c/em\u003e = -0.11) and PDW (\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.21). MgSO4 is effective as a replacement for K2-EDTA in dogs. MgSO4 in cats is critical because it helps prevent platelet clumping, thereby maintaining normal PLT and PCT levels. The beneficial compound in feline blood analysis requires caution in use. There are no favorable MgSO4 values for K2-EDTA with respect to the size parameters MPV and PDW. It suggests such parameters are not consistently accurate, irrespective of the anticoagulant used.\u003c/p\u003e","manuscriptTitle":"Assessing the analytical reliability of platelet counts and related indices in dogs and cats: a correlation matrix analysis comparing K2-EDTA and MgSO4 anticoagulation","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-02-26 18:42:47","doi":"10.21203/rs.3.rs-8282197/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"60cd9931-22af-4154-bd9e-2e9a230bb399","owner":[],"postedDate":"February 26th, 2026","published":true,"recentEditorialEvents":[{"type":"decision","content":"Rejected","date":"2026-05-12T12:06:31+00:00","index":"","fulltext":""}],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-05-12T13:15:00+00:00","versionOfRecord":[],"versionCreatedAt":"2026-02-26 18:42:47","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8282197","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8282197","identity":"rs-8282197","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}