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Comparing to the LDPE membrane this was a reduction of dialysis time from 160 to 120 min, but a higher solvent use of 150 ml per sample.
Blood samples (approximately 0.2 mL per sample) were collected from each animal via jugular vein cannulae at the following time points: predose; 5, 15, and 30 min post dose; and 1, 2, 4, 8, and 24 h post dose.
The analytical procedure performed with PLE showed similar recoveries (∼70%) and total PAH concentrations (∼200 ng g−1) as found using Soxtec extraction, with the advantage of reducing solvent consumption by 3 (30 mL against 100 mL per sample), and taking a fifth of the time (24 samples extracted automatically in 8 h against 2 samples in 3.5 h).
A volume of 5 ml per sample was filled in a dry and clean sample cup (Ø 40 mm, 12 mm, polypropylene, ePW sample cups, Novasina) and each sample was treated 10 min in the pre-condition chamber of the instrument before measurement.
A lytic enzyme cocktail was prepared at the time of extraction and added to each sample as follows: 50 µl Lysozyme (450 kU ml-1), 6 µl Mutanolysin (25 kU ml−1), 3 µl Lysostaphin (4 kU ml−1) and 41 µl TE50 for a final volume of 100 ml per sample.
In the case of the MEPS procedure, waste production is only about 1.5 mL per sample.
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Sample volumes were 100 ml per sampling point, the samples were then stored at temperatures not higher than 4 °C and were analyzed within 2 months of sampling.
Peripheral blood samples (6 ml per sampling time) were collected in heparinised tubes and centrifuged at 1000 g for 15 min at 4°C.
According to the manufacturer's protocol 1 ml urine per sample was lysed and RNA was isolated and purified in a spin column procedure.
Briefly, a 24-well head is used to isolate 1.8 mL of blood per sample.
Tumor samples from patients with residual disease were also collected at definitive surgery.Serum samples (4.9 mL whole blood per sample) for biomarker assessment were collected at baseline and at the time of definitive surgery.
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