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To further evaluate the coordinated action of ProDH with P5CDH or P5CR, we quantified the content of Pro and its derivatives at each HR stage.
The LV was calculated using equations 2 and 3 at each HR using the measured width FWHM and depth D. The data are plotted in ESM Figs. 7 and 8 for the longitudinal and axial plane, respectively, and have been fitted to a line.
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The number of dead animals and the number of total animals were counted to determine percent survival of mua-3 and wild-type at each hour from 50 hr at 15°, 37 hr at 20°, and 30 hr at 25°.
After fixation, guts were washed in antibody wash with continuous shaking three times for at least 20 min followed by primary and secondary antibody application for at least 12 hr each at 4°C with at least 12 hr of washing with antibody wash in between.
After clipping and at each time point (0 hr, 6 hr, 24 hr) before collecting the biopsy samples, skin was evaluated for signs of inflammation.
At 0800 hr each day, before the introduction of fresh subsamples of the effluent, water/effluent samples (700 mL each) were removed from each exposure tank (and from the effluent holding tank) and extracted onto primed C18 solid-phase extraction columns for assessment of estrogenic activity in the rYES, following the procedures described above.
At each time point, HR and SpO2 were measured using the three connected devices and the gold standard.
For the 5HT-induced and original varicosities, the number of varicosities in each category for each coculture at 48 hr was normalized to the number in the same category in that coculture at 24 hr.
Exhaled breath was collected in a Mylar balloon at approximately 1600 hr each day using an offline F eNO protocol.
Dissected tissues were fixed in 4% paraformaldehyde (PFA) and cryoprotected by incubation in a sucrose gradient for at least 1 hr each (5%, 15 %, 30).
This is further substantiated by the fact that after approximately 2 years of an antipsychotic-free follow-up period for each individual at HR in this sample, only 35(5%) made a transition to a psychotic disorder.
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