An assessment of a biosensor system for the quantification of microcystins in freshwater cyanobacterial blooms.

Microcystin-producing cyanobacterial blooms are a global issue threatening drinking water supplies and recreation on lakes and beaches. Direct measurement of microcystins is the only way to ensure waters have concentrations below guideline concentrations; however, analyzing water for microcystins takes several hours to days to obtain data. We tested LightDeck Diagnostics' bead beater cell lysis and two versions of the quantification system designed to give microcystin concentrations within 20 min and compared it to the standard freeze-thaw cycle lysis method and ELISA quantification. The bead beater lyser was only 30 % effective at extracting microcystins compared to freeze-thaw. When considering freeze-thaw samples analyzed in 2021, there was good agreement between ELISA and LightDeck version 2 (n = 152; R2 = 0.868), but the LightDeck slightly underestimated microcystins (slope of 0.862). However, we found poor relationships between LightDeck version 2 and ELISA in 2022 (n = 49, slopes 0.60 to 1.6; R2 < 0.6) and LightDeck version 1 (slope = 1.77 but also a high number of less than quantifiable concentrations). After the quantification issues are resolved, combining the LightDeck system with an already-proven rapid lysis method (such as microwaving) will allow beach managers and water treatment operators to make quicker, well-informed decisions.

Clickable decellularized extracellular matrix as a new tool for building hybrid-hydrogels to model chronic fibrotic diseases in vitro.

Fibrotic disorders account for over one third of mortalities worldwide. Despite great efforts to study the cellular and molecular processes underlying fibrosis, there are currently few effective therapies. Dual-stage polymerization reactions are an innovative tool for recreating heterogeneous increases in extracellular matrix (ECM) modulus, a hallmark of fibrotic diseases in vivo. Here, we present a clickable decellularized ECM (dECM) crosslinker incorporated into a dynamically responsive poly(ethylene glycol)-α-methacrylate (PEGαMA) hybrid-hydrogel to recreate ECM remodeling in vitro. An off-stoichiometry thiol-ene Michael addition between PEGαMA (8-arm, 10 kg mol-1) and the clickable dECM resulted in hydrogels with an elastic modulus of E = 3.6 ± 0.24 kPa, approximating healthy lung tissue (1-5 kPa). Next, residual αMA groups were reacted via a photo-initiated homopolymerization to increase modulus values to fibrotic levels (E = 13.4 ± 0.82 kPa) in situ. Hydrogels with increased elastic moduli, mimicking fibrotic ECM, induced a significant increase in the expression of myofibroblast transgenes. The proportion of primary fibroblasts from dual-reporter mouse lungs expressing collagen 1a1 and alpha-smooth muscle actin increased by approximately 60% when cultured on stiff and dynamically stiffened hybrid-hydrogels compared to soft. Likewise, fibroblasts expressed significantly increased levels of the collagen 1a1 transgene on stiff regions of spatially patterned hybrid-hydrogels compared to the soft areas. Collectively, these results indicate that hybrid-hydrogels are a new tool that can be implemented to spatiotemporally induce a phenotypic transition in primary murine fibroblasts in vitro.