Of the latter 70 samples, 18% were excluded because of poor quality (excessive debris, large percentage of epithelial cells present), 16% were excluded because of a recovered volume < 20 mL and 66% were excluded because of a low total cell count (< 60,000 cells/mL).
One ml of the recovered volume was used for bacterial culture, the other part (2 3 ml) was diluted with 20 ml of phosphate buffered saline (PBS), then homogenized and centrifuged (400 g for 10 minutes) to gain cell pellet and supernatant.
We did not monitor recovered volumes but used an instillation volume of 150 ml for our study, which is at the lower end of recommended values.
Recovered volumes were measured before aliquoting.
Peaks were quantified by area integration and the concentrations were normalized to the quantity of the internal standard recovered, and volume of sample processed, derivatized, and injected [ 75].
The final sample of 50 100 µg of sVSG221 was recovered in a volume of 100 µL water.
The resulting probes were purified with a Microcon microcentrifuge filter device and recovered in a volume of 25 μl.
Aerated tap water containing the same Zn concentration was subsequently added to recover the initial volume of the aquaria.
