A Robust Chemical-free Platform for Age-Synchronized Caenorhabditis elegans Populations Maintenance for High-Throughput Screening in Aging Studies

Abstract Caenorhabditis elegans (C. elegans) is a well-established model for investigating the mechanisms of aging and age-related disorders like neurodegeneration. However, maintaining age-synchronized populations of C. elegans for aging studies without genetic or pharmacological interventions presents significant challenges, given their high reproductive rate: each nematode produces over 300 progeny. The traditional method for maintaining an age-synchronized population for multi-day studies without interventions is labor-intensive and low-throughput, hindering research on aging mechanisms and the identification of novel interventions for aging. To address these limitations, a novel, robust method was developed to sustain age-synchronized populations in a 96-well plate liquid culture format for up to 12 days without custom-made apparatuses. The robustness of this method was substantially improved by optimizing the surface composition of the multi-well plate and disposables, refining culture parameters, including life stage, medium composition, and bacterial food concentration. To facilitate unbiased phenotype assessment throughout the lifespan, we used a Wmicrotracker ONE reader to monitor worm movement and viability in a multi-well plate. The overall fitness decline with aging using our method is comparable to that of worms maintained on solid agar. Lastly, using transgenic C. elegans carrying tauopathy, we demonstrated the ability of applying our optimized platform for high-throughput screening with a Z-factor of 0.7. Our novel method simplifies age-synchronized population maintenance, enhances progeny separation, and reduces costs, enabling high-throughput screening of compounds and RNAi libraries. These advancements greatly enhance the versatility of C. elegans as a model organism, offering a scalable platform for genetic and compound screening and for comprehensive investigations into drug discovery and disease mechanisms. Competing Interest Statement The authors have declared no competing interest.