For this purpose, the production medium was dispensed into flasks of 250 mL capacity; each flask containing 50 mL of liquid medium was adjusted at pH 6.5.
Diluted aliquots (∼4 × 10 cells in 50 μl of the same medium) were dispensed into wells of a 96-well plate containing 50 μl of the same medium or 50 μl of a twofold dilution series of GE in the same medium (final concentrations = 0 and 8 8000 μg/ml), and were incubated 16 hr at 37°C with shaking under the same conditions.
The medium was dispensed into the culture tubes while being gassed with 100% CO2, at 9.5 ml of medium per tube, and the tubes sealed with blue butyl septum stoppers and aluminium seals (Bellco), with a headspace of 100% CO2.
In addition, the prepared enriched " Trichomonas medium" was dispensed aseptically into sterile McCartney bottles in 10 ml aliquots and used as both transport and culture medium for all the samples collected for isolation of T. foetus with microscopic examination of the medium at intervals from day 1 to day 7 after inoculation, taken from the bottom of the bottle.
Unless specified otherwise, cells (5 × 10) were suspended in 0.1 ml of 0.1% albumin-containing culture medium and dispensed into the upper chambers of Transwells (8 μm pore size), with 0.6 ml of 10% FBS-containing medium in the lower chamber.
The constituents of 50 mL of this medium were dispensed in flasks of 250 mL capacity.
Under aseptic conditions in the laminar flow hood, the medium was dispensed into 150 mm pre-sterilized petri dishes to yield a uniform depth of 4 mm.
According to the experimental protocols, 6 ml of culture samples were periodically withdrawn at 0, 2, 4 and 6 h of cultivation, centrifuged 10 min at 12,000 rpm, resuspended in 600 μl by using their own medium and dispensed in three wells.
The biofilm formation was carried out using the P63 strain of Pseudomonas aeruginosa and the M63 minimal medium [M63 salt 12 g/l KH2PO4, 28 g/l K2HPO4, 8 g/l (NH4) SO4, 1 mM MgSO4, 0.5%] 150 μl of M63 liquid medium was dispensed into two 12-well plates, and the seed culture of the clinical cultures cultured for 24 h was inoculated at 2% and cultured at 30 °C for 72 h.
