PNNL

Abstract

Biologically derived neurotoxins from cyanobacteria and algae impact environmental resources in addition to being considered a potential biological threat to human and animal health. Activity-based assays are essential to detecting and responding to toxic neurotoxin events either naturally occurring or deliberate. Two toxins of interest include saxitoxin and brevetoxin. These toxins bind to and alter the function of voltage-gated sodium channels (NaV channels) which are essential for generating cell membrane action potential. We report the development and refinement of a System for Analysis at Liquid Vacuum Interface (SALVI) to assess the functional activity of saxitoxin and brevetoxin. This approach utilizes a vacuum-compatible microfluidic reactor that permits analysis at the liquid vacuum interface of human derived cells with a neurotoxin of interest in a biologically relevant environment. SALI based detection is technically relevant because of the following: • Compatible with multiple detection modalities for real-time monitoring o Optical imaging techniques o Mass spectrometry characterization via ToF-SIMS • Flexible platform that can be modified: o Numerous biological tissues can be integrated into the system o Types of marine toxin to be evaluated This LDRD effort improved technical methods resulting in a reproducible mammalian cell cultivation and neurotoxin exposure system to detect differences of chemical signatures within the SALVI system. Additionally, this effort resulted in the following capability development: • This project involved cross directorate collaborations (NSD and EBSD) to build cross-disciplinary capabilities that did not previously exist at PNNL. This involved cross-directorate networking and expanding microfluidics capabilities for national security research in Chemical and Biological Science group. The project also utilized the ToF-SIMS in EMSL for all measurements. • Strategic support of multi-directorate PNNL staff members and refined mechanical and instrument operation analysis with ToF-SIMS. • Methodology for the improved development of a SALVI prototype and assay to detect chemical biomarkers of neurotoxins in aquatic environments with a non-commercial system that offers flexibility for future efforts. Future efforts will enable improved SALVI protocol development for multiple human tissue types and aquatic toxins. Additionally, multiple detection modalities could be implemented for enhanced confidence across an array of biological threat characterization measurements.