We engineered yeast strains containing fluorescently labeled peroxisomes and lacking either Dnm1, Vps1, or Fis1, cultured these cells in oleic acid medium, counted the number of peroxisomes in each cell, and calculated the mean and Fano factor of the peroxisome abundance distributions for each strain.
On the final day of culture anti-CD3/CD28 mAb-coated beads were removed, cells were washed in medium, counted in the presence of a trypan blue dye and resuspended in medium containing 50 IU/mL of IL-2 for 24 48 h prior to all phenotypic or functional assays.
Resuspend the pellet in 1 mL of medium, count cells and seed the passaged PBECs onto Transwell inserts at 1.0×10 cells/cm.
SKOV3ip cells were detached with trypsin/EDTA, resuspended in culture medium, counted and divided into aliquots of equal cell number.
Resuspend the pellet in 1 mL of basic growth medium, count cells and seed onto Transwell inserts at 8×10 cells/insert.
The cells were resuspended with 10 ml of primary hepatocyte medium, counted, and plated in 12-well plates (coated with 0.2% gelatin) at a density of 2 × 10 cells/well.
These C. albicans cells were washed with complete culture medium, counted with a hemocytometer and diluted to 5.0 × 106 cell/ml in 50 ml of complete culture medium.
Once isolated on a solid medium, the counting of Bc-like colonies appears to be a reproducible method for the quantification of the bacterial population from our samples.
In this present study, the initial bacterial load (7.12 ± 0.12log10CFU/lung) was higher than a medium inoculum count of 5*10 CFU/ml routinely used in time-kill studies.
A counting frame of 25 μm × 25 μm and a counting grid of 85 μm × 85 μm was used to sample regions throughout the nucleus; counts of medium to darkly labeled cells were made using standard stereological rules, as previously described [ 36].
