Refinement of an open-microcavity optical biosensor for bacterial endotoxin test.

The Limulus Amebocyte Lysate (LAL) test is an in vitro assay widely used in the pharmaceutical and biotechnology industries to detect bacterial endotoxins. Endotoxin is a structural component of the cell wall of Gram-negative bacteria, which has serious pathogenic effects in the body and may cause dysfunction of multiple organ systems and increased risk of mortality. To address the growing need for LAL assays due to the increased demand from drug and vaccine manufacturers, we have developed a new LAL assay approach. Our detection mechanism is different and improved from those currently used in the industry, leading to increased test sensitivity and reduced assay time. Our study utilizes an open-microcavity photonic-crystal biosensor to quantify endotoxin concentrations. It has demonstrated an improved LAL assay sensitivity by 10 fold compared to the commercial standard methods and reduced the time needed for the assay by more than half. In addition, this approach requires as little as 5 µL of LAL reagent per test, thereby decreasing costs and conserving horseshoe crabs. The results reported in this paper indicate the possibility of using the photonic-crystal biosensor based approach for significant enhancements of endotoxin testing.

Study of the Interactions of Fusarium virguliforme Toxin FvTox1 with Synthetic Peptides by Molecular Simulations and a Label-Free Biosensor.

Fusarium virguliforme is a soil borne pathogen that causes sudden death syndrome (SDS) in soybean plants. This pathogenic disease may result in severe soybean yield suppression and can cause serious economic harm. It has been shown that the FvTox1 toxin produced by the pathogen may be the root cause of foliar SDS. Anti-FvTox1 single-chain variable fragment antibody expressed in transgenic soybean plants was shown to neutralize the FvTox1 toxin involved in foliar SDS development. Here, we have investigated the binding affinities of FvTox1 with four FvTox1-interacting peptides of 7 to 12 amino acids identified from phage display libraries using both bioinformatics-based molecular simulations and label-free bioassays with a unique photonic crystal biosensor. Results from the molecular simulations have predicted the interaction energies and 3-dimensional (3D) structures of FvTox1 and FvTox1-interacting peptide complexes. Our label-free binding assays have further provided the interaction strength of FvTox1 with four different FvTox1-interacting peptides and experimentally confirmed the simulation results obtained from bioinformatics-based molecular calculations.