Paper-Based Analytical Device for Real-Time Monitoring of Egg Hatching in the Model Nematode Caenorhabditis elegans.

ABSTRACT

Caenorhabditis elegans is an in vivo model known for its easy handling and maintenance and lack of associated ethical issues. The release of chitinase can be used to monitor the egg-laying stage in C. elegans. The aim of this study was to develop a simple and cost-effective device to monitor the activity of chitinase in embryos of C. elegans. Colloid chitin azure (CCA), a substrate for chitinase, was preimmobilized on the detection area of paper, forming a purple region, to generate a CCA paper-based analytical device (CCA-PAD). The degradation of CCA by chitinase could be observed as the purple color became faint and the filter paper eventually became colorless. Under the optimum conditions, the proposed device quantified the chitinase enzyme in the range of 15.625-125 mU/mL within 48 h (R2 = 0.993). In this work, 10 young adult-staged wild-type C. elegans (Bristol N2) worms were analyzed on the CCA-PAD, which was supplemented with the laboratory food source E. coli OP50 on a gauze layer. The same strain treated with 5-fluoro-2'-deoxyuridine was used to prevent egg production in C. elegans. A significant difference in the color intensity was observed between these two groups at the end of the experiment (P = <0.001, independent t-test, n = 3). We successfully developed a simple and effective method for monitoring chitinase activity. The device may have potential applications in drug-screening studies as it efficiently distinguishes drugs that can impact egg laying.