Application of a Modified Culturing Method for Lymphocytes in Cytogenetic Research

Objective To establish a modified method for microculturing whole human blood for cytogenetic analysis.Methods A novel tube rack was designed to overcome the drawbacks of directly culturing the cells within centrifuge tubes. The fractions of human plasma, human serum and two commercial fetal bovine sera were analyzed with 1 5% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).The influence of adding 0%, 5%, 10%, 1 5%,20%,25% and 30%autologous plasma to the culture on lymphocyte transformation rate and mitotic index (MI)was examined. Results The SDS-PAGE analysis showed a significant difference between commercial fetal bovine sera,and that the components of human plasma were similar to those of fetal bovine serum.The value of MI in lymphocyte was evidently increased along with addition of autologous plasma.However,this has exerted no significant effect on the transformation rate.With the addition of 10% autologous plasma,the MI value has become much higher than the conventional method.Conclusion A modified method was established by application of a novel tube inclined rack and optimization of whole blood inoculation.This method is easier and cheaper,and is suitable for application in clinical practice.