Optimization, Application, and Interpretation of Lactate Dehydrogenase Measurements in Microwell Determination of Cell Number and Toxicity

The lactate dehydrogenase (LDH) assay was addressed for its sensitivity, disturbances by foaming, and cell number and size. Cells were from a U-251 MG grade IV human glioblastoma brain tumor cell line used in 100-µl well volumes. Cells were counted by microscopy and Coulter counting; assays were LDH or trypan blue. The results indicate increased 490 nm signals (level, variance) by using phenol red or by increasing fetal bovine serum from 5% to 10%. The data also indicate that defoaming results in reduced variances ranging from a factor of 2 at 1–3 units of absorption, up to a factor of 4–5 at