Collagenase and Dispase enzymes disrupt lymphocyte surface molecules

Collagenase and Dispase enzymes are often used to disaggregate tissue. In this study, we examined by flow cytometry the effects of these enzyme preparations on peripheral blood lymphocyte surface marker expression. Peripheral blood mononuclear cells (PBMC) were obtained from seven healthy volunteers. Cells were incubated with collagenase (128 U/ml, type 1 A) for 3 h and overnight and with Dispase (1.6 mg/ml, grade II) for 15 min, 30 min, 1 h, 3 h and overnight. The intensity of expression of CD3, CD4, CD8, alpha beta and gamma delta T cell receptors was decreased by 25-40% in all cases, while CD4+ and CD8+ lymphocyte populations were undetectable after treatment with Dispase. Moreover, reappearance of these surface molecules did not occur following the incubation of cells in culture medium for 3 h. The results of this study emphasise the importance of considering the influence of isolation procedures on cell characteristics.