Direct and Rapid Measurement of Hydrogen Peroxide in Human Blood Using a Microfluidic Device

The levels of hydrogen peroxide (H 2 O 2 ) in human blood is of great relevance as it has emerged as an important signalling molecule in a variety of disease states. Fast and reliable measurement of H 2 O 2 levels in the blood, however, continues to remain a challenge. Herein we report an automated method employing a microfluidic device for direct and rapid measurement of H 2 O 2 in human blood based on laser-induced fluorescence measurement. Our study delineates the critical factors that affect measurement accuracy – we found blood cells and soluble proteins significantly alter the native H 2 O 2 levels in the time interval between sample withdrawal and detection. We show that separation of blood cells and subsequent dilution of the plasma with a buffer at a ratio of 1:6 inhibits the above effect, leading to reliable measurements. We demonstrate rapid measurement of in plasma in the concentration range of 0 – 49 µM, offering a limit of detection of 0.05 µM, a sensitivity of 0.60 µM -1 , and detection time of 15 min; the device is amenable to the real-time measurement of H 2 O 2 in the patient’s blood. Using the linear correlation obtained with known quantities of H 2 O 2 , the endogenous H 2 O 2 concentration in the blood of healthy individuals is found to be in the range 2 – 6 µM. The availability of this device at the point of care will have relevance in understanding the role of H 2 O 2 in health and disease.