A simple method to predict cellular density in adipocyte metabolic incubations

OBJECTIVES: The density of isolated adipocyte suspensions, namely the cellular concentration, influences metabolic results when lipolysis and the pattern of glucose metabolism are studied. It is often difficult to obtain reproducible adipocyte concentrations from experiment to experiment, and investigators usually measure the cell concentration at the end rather than at the initiation of metabolic incubations. METHOD: A simple and rapid method to obtain reliable and predictable adipocyte concentration prior to metabolic incubations is described and validated. The method is based on determination of lipocrit, mean adipocyte diameter (by optical sizing), and calculation of volume, in aliquots of isolated adipocyte suspensions. MAIN OUTCOME MEASURES: Lipocrit, mean adipocyte volume, predicted and observed adipocyte number in isolated cell suspensions. RESULTS: In 15 experiments, adipocyte concentration was accurately predicted within 12-18% of actual concentration. This is in contrast to the four-or five-fold differences usually encountered in a series of experiments. CONCLUSION: One can rapidly predict the number of adipocytes present in a given cell suspension with the proposed method, and then correct it to a desired adipocyte concentration at the beginning of metabolic incubations. This method will help to eliminate the confounding effects of variable cell concentrations in in vitro metabolic experiments with isolated adipocytes.