Multiplex Lateral Flow Assay and the Sample Preparation Method for the Simultaneous Detection of Three Marine Toxins.

A multiplex lateral flow immunoassay (LFA) has been developed to detect the primary marine biotoxin groups: amnesic shellfish poisoning toxins, paralytic shellfish poisoning toxins, and diarrhetic shellfish poisoning toxins. The performance characteristics of the multiplex LFA were evaluated for its suitability as a screening method for the detection of toxins in shellfish. The marine toxin-specific antibodies were class-specific, and there was no cross-reactivity between the three toxin groups. The test is capable of detecting all three marine toxin groups, with working ranges of 0.2-1.5, 2.5-65.0, and 8.2-140.3 ng/mL for okadaic acid, saxitoxin, and domoic acid, respectively. This allows the multiplex LFA to detect all three toxin groups at the EU regulatory limits, with a single sample extraction method and dilution volume. No matrix effects were observed on the performance of the LFA with mussel samples spiked with toxins. The developed LFA uses a simple and pocket-sized, portable Cube Reader to provide an accurate result. We also evaluated the use of this Cube Reader with commercially available monoplex lateral flow assays for marine toxins.

High-Throughput Time-Lapse Fluorescence Microscopy Screening for Heterogeneously Expressed Genes in Bacillus subtilis.

Elucidating phenotypic heterogeneity in clonal bacterial populations is important for both the fundamental understanding of bacterial behavior and the synthetic engineering of bacteria in biotechnology. In this study, we present and validate a high-throughput and high-resolution time-lapse fluorescence microscopy-based strategy to easily and systematically screen for heterogeneously expressed genes in the Bacillus subtilis model bacterium. This screen allows detection of expression patterns at high spatial and temporal resolution, which often escape detection by other approaches, and can readily be extrapolated to other bacteria. A proof-of-concept screening in B. subtilis revealed both recognized and yet unrecognized heterogeneously expressed genes, thereby validating the approach. IMPORTANCE Differential gene expression among isogenic siblings often leads to phenotypic heterogeneity and the emergence of complex social behavior and functional capacities within clonal bacterial populations. Despite the importance of such features for both the fundamental understanding and synthetic engineering of bacterial behavior, approaches to systematically map such population heterogeneity are scarce. In this context, we have elaborated a new time-lapse fluorescence microscopy-based strategy to easily and systematically screen for such heterogeneously expressed genes in bacteria with high resolution and throughput. A proof-of-concept screening in the Bacillus subtilis model bacterium revealed both recognized and yet unrecognized heterogeneously expressed genes, thereby validating our approach.