Determination of cell numbers and viability; comparison of the Microcyte flow cytometer with trypan blue counts and coulter counts

摘要

Traditionally in mammalian cell culture the amount of biomass in a culture is expressed as a cell density. Cell densities are commonly measured by manually counting dilutions of a culture in the presence of a staining agent like trypan blue. For larger numbers of samples this method is not only time consuming but also prone to operator dependent variations. Nielsen et al. have shown that variability between individuals and counting of relatively small numbers of cells both contribute to a large margin of error in manual trypan blue counts. Instrument mediated ways of cell counting can lead to a higher standardization of counting results and if a large number of cells is processed within one measurement the margins of error will also improve. To yield the same information as a manual count of a trypan blue stained cell sample the instrument mediated method should be able to distinguish between viable and non-viable cells. Recently a compact portable flow cytometer was marketed which can be used for cell enumeration. The use of the fluorescent nucleic acid stain TOPRO-3 allows simultaneous determination of total cell number and the number of non-viable cells. The measuring principle of the Microcyte flow cytometer is shown in Fig. 1.