Rapid analysis of lymphocyte subsets in cord blood.

Several investigators have enumerated cellular populations in neonatal cord blood with variable results. In this study, the authors established reference ranges for lymphocyte subsets in cord blood from healthy newborns using a whole blood lysis technique on the Coulter Immunoprep Epics Leukocyte Preparation System (Coulter Immunology, Hialeah, FL). All analyses were performed on a flow cytometer by gating on forward angle versus 90 degrees light scatter. Lymphocytes demonstrated all surface markers examined, including T4, T8, T3, T11, B1, NKH-1, I3, and 4B4; 2H4 suppressor inducer lymphocytes were prominent in neonatal blood. The authors think this standardized system may be suitable for use in neonatal and pediatric patients because it quickly processes small aliquots of whole blood with minimal sample manipulation.

Production of monoclonal antibody against Pneumocystis carinii by using a hybrid of rat spleen and mouse myeloma cells.

It has been difficult to obtain pure Pneumocystis carinii antigen either from cultures or from infected lungs for use in producing a specific antibody against P. carinii. This report describes an approach toward producing a monoclonal antibody that bypasses the antigen purification steps. P. carinii infection was developed in Sprague-Dawley rats by the method of immunosuppression with cortisone. The infected lungs were homogenized, and the homogenate was used to immunize Sprague-Dawley rats. Rat spleen cells were then fused with SP2/0 mouse myeloma cells. Hybridoma clones were screened for antibody production against P. carinii by immunoperoxidase staining techniques and by enzyme-linked immunosorbent assay, using as antigens homogenates of normal rat lung, homogenates of P. carinii-infected rat lung, and harvests of P. carinii grown with WI-38 cells. Out of six hybridoma clones obtained that produced antibodies against P. carinii, one was able to produce ascitic fluid. This monoclonal antibody reacted with two P. carinii antigens with masses of about 35,000 and 65,000 daltons in P. carinii-infected lungs and three proteins with masses of about 35,000, 65,000, and 110,000 daltons in P. carinii that was harvested from a WI-38 cell culture.