Long-term storage and impedance-based water toxicity testing capabilities of fluidic biochips seeded with RTgill-W1 cells

Rainbow trout gill epithelial cells (RTgill-W1) are used in a cell-based biosensor that can respond within one hour to toxic chemicals that have the potential to contaminate drinking water supplies. RTgill-W1 cells seeded on enclosed fluidic biochips and monitored using electric cell-substrate impedance sensing (ECIS) technology responded to 18 out of the 18 toxic chemicals tested within one hour of exposure. Nine of these chemical responses were within established concentration ranges specified by the U.S. Army for comparison of toxicity sensors for field application. The RTgill-W1 cells remain viable on the biochips at ambient carbon dioxide levels at 6°C for 78 weeks without media changes. RTgill-W1 biochips stored in this manner were challenged with 9.4 μM sodium pentachlorophenate (PCP), a benchmark toxicant, and impedance responses were significant (p \u3c 0.001) for all storage times tested. This poikilothermic cell line has toxicant sensitivity comparable to a mammalian cell line (bovine lung microvessel endothelial cells (BLMVECs)) that was tested on fluidic biochips with the same chemicals. In order to remain viable, the BLMVEC biochips required media replenishments 3 times per week while being maintained at 37°C. The ability of RTgill-W1 biochips to maintain monolayer integrity without media replenishments for 78 weeks, combined with their chemical sensitivity and rapid response time, make them excellent candidates for use in low cost, maintenance-free field-portable biosensors

Zebrafish morphological endpoints.

This table shows all the endpoints that were recorded for each test at 30hpf (early morphological endpoints) and 120hpf (late morphological endpoints). Abbreviations for the endpoints are on the left side of the table with the corresponding descriptor on the right side. (DOCX)</p

Table 1 -

Table 1 - </p

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Table 2 - </p

Mortality curves of zebrafish embryos exposed to different nanomaterials.

120 hpf mortality curves of zebrafish embryos exposed to different nanomaterials. Mortality curves consist of multiple tests. If the concentrations between tests of the same nanomaterial overlapped, the average mortality at the concentration was reported in the mortality curve. This figure shows the mortality curves of silver nanomaterials with the concentrations adjusted for the concentration of ionic silver in solution. (120 hpf (LC50) for the silver nanomaterials with adjusted concentrations are as followed: silver nitrate (0.08 μg/mL), 5 nm nano silver (0.03 μg/mL), 25 nm nano silver (0.04 μg/mL), and 75 nm nano silver (0.16 μg/mL)).</p

Mortality curves of zebrafish embryos exposed to different nanomaterials.

120 hpf mortality curves of zebrafish embryos exposed to different nanomaterials. Mortality curves consist of multiple tests. If the concentrations between tests of the same nanomaterial overlapped, the average mortality at the concentration was reported in the mortality curve. This figure shows the mortality curve of silver nanomaterials based on size without the concentrations being adjusted for the ionic concentration of silver in solution. (120 hpf (LC50) for the silver nanomaterials are as followed: silver nitrate (0.08 μg/mL), 5 nm nano silver (0.71 μg/mL), 25 nm nano silver (3.68 μg/mL), 75 nm nano silver (33.1 μg/mL), and 75 nm nano silver (NIST) (43.3 μg/mL)).</p

Time series test for the photomotor response of zebrafish embryos.

Zebrafish photomotor response by phase. The graph depicts the control response of zebrafish embryos (n = 603) with increasing developmental age. Data generated from the PMR assay was further condensed using the area under the curve calculation by phase with standard deviation bars. Embryos that were found to be non-responders were excluded from the dataset. * = statistically significant increase in movement during the excitatory phase over the background phase.</p