An inexpensive, high-throughput μPAD assay of microbial growth rate and motility on solid surfaces using Saccharomyces cerevisiae and Escherichia coli as model organisms

Abstract

Many microbial phenotypes are differentially or exclusively expressed on agar surfaces, including biofilms, motility, and sociality. However, agar-based assays are limited by their low throughput, which increases costs, lab waste, space requirements, and the time required to conduct experiments. Here, we demonstrate the use of wax-printed microfluidic paper-based analytical devices (μPADs) to measure linear growth rate of microbes on an agar growth media as a means of circumventing the aforementioned limitations. The main production materials of the proposed μPAD design are a wax printer, filter paper, and empty pipet boxes. A single wax-printed μPAD allowing 8 independent, agar-grown colonies costs $0.07, compared to $0.20 and $9.37 for the same number of replicates on traditional microtiter/spectrophotometry and Petri dish assays, respectively. We optimized the μPAD design for channel width (3 mm), agar volume (780 μL/channel), and microbe inoculation method (razor-blade). Comparative analyses of the traditional and proposed μPAD methods for measuring growth rate of nonmotile (Saccharomyces cerevisiae) and motile (flagellated Escherichia coli) microorganisms suggested the μPAD assays conferred a comparable degree of accuracy and reliability to growth rate measurements as their traditional counterparts. We substantiated this claim with strong, positive correlations between the traditional and μPAD assay, a significant nonzero slope in the model relating the two assays, a nonsignificant difference between the relative standard errors of the two techniques, and an analysis of inter-device reliability. Therefore, μPAD designs merit consideration for the development of enhanced-throughput, low-cost microbial growth and motility assays.

Efficient and Simple Paper-Based Assay for Plasma Separation Using Universal Anti-H Agglutinating Antibody

“…Following that, a wood burner device was used to melt the wax onto the filter paper to draw the required channels . The second method was waxing channels automatically using a Xerox 8580 solid ink printer, then melting the wax from the surface of the filter paper into its layers using a laminator device . Different filter papers were chosen for the membrane filtration approach (Whatman Qualitative filter paper grades 5 (2.5 μm) and Whatman Quantitative ashless filter papers grade 42 and 44 (2.5 and 3 μm, respectively)).…”