Flow cytometry for assessment of white blood cells subsets in standard buffy coat product

Abstract

Circulating white blood cells (WBC) play an important role in the human immune system including both the innate and adaptive immunity. The immune cells such as granulocytes (neutrophils, eosinophils, basophils), lymphocytes, and monocytes are regularly found in blood components known as Buffy Coat. Most of the time the buffy coat serves no clinical purpose since the concentrated immune cells (especially the lymphocytes) can trigger the transfusion-associated graft versus the host diseases (TA-GVHD). In special circumstances, such as neutropenia patients undergoing chemotherapy or hematopoietic stem cell transplant (HSCT) with uncontrolled infection, patients require granulocytes transfusion to compensate for the deficiency in the body defense system. However, many of in vitro research studies require the use of peripheral blood mononuclear cell (PBMC) for the study of the human immune system. This can be made possible by isolating the buffy coat. But the exact proportion of various immune cells within the buffy coat especially among Chinese donors remains unknown. In this study, about 30 healthy adult blood donors between 22 and 60 years of age were recruited to establish the reference value of the white blood cells subsets from the buffy coat as defined by the flow cytometer analysis. The 5 monoclonal antibodies of CD 45-PB, CD16-APC-Cy7, CD14-PE, CD19-APC, and CD3-FTIC were used to identify the panleukocytes, neutrophils, monocytes, B- lymphocytes, and T-lymphocytes respectively. My result demonstrated the differences in the number of the white blood cells subtypes between gender, such as the female donors had a significantly lower CD16+ neutrophils and a higher CD3+ T-lymphocytes,. In this study, it can be concluded that the neutrophils are very fragile, and difficult to maintain their integrity and survival in the unfavorable storage condition. However it could still be demonstrated within the white blood cells subsets. In the future, higher sample size can be investigated to verify the gender effects on the white blood cells subpopulation.