For selected experiments, iodo acetate was added by sterile filtration to the sterilized growth medium giving a final concentration of 1 mM.
When HOFs and HOKs were 70 80% confluent, dye containing polymersomes were added to the cell culture medium giving a final concentration of 1 mg polymer/ml cell culture media.
Analysis of the fermentations with CM1 medium gave a similar difference in acid ratio production (Fig. 3b).
(19) The liquid crystalline medium gave a stable quadrupolar splitting of the D2O H signal of 29.9 Hz, with a final concentration of apo-DHFR of ∼0.21 mM.
Glucose was added to the medium giving a final concentration of 20 g l-1 taken into account the amount of glucose already present in the hydrolysate.
Bromocresol Purple was used to indicate the pH of the medium, giving a color shift, from yellow to purple, during the acidification of the liquid medium: this change correlated well with the growth of the yeast cells [ 14].
The medium was replaced and 100 μl transfection mixture containing 3 μl Lipofectamine 2000 (Invitrogen, Carlsbad, Ca) and 3 μl 20 μM LPAR3 ON-TARGET plus siRNA (Dharmacon, Lafayette, CO) in OptiMEM was added pr well (1 ml medium), giving a final concentration of 60 nM siRNA.
Aliquots of the virus/drug suspension were then diluted 500 fold and plated in triplicate on aspirated HeLa, A549 and SIRC cell monolayers to initiate infection (6 8 × 105 cells/cm2 in T-25 culture flasks containing 9.9 mls fresh culture medium giving a total volume of 10 mls with an MOI of 10).
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