Isolation of Ia positive human leukocytes by a direct rosette assay.
A direct rosette assay (DRA) using ox erythrocytes coated with monoclonal antibody (AF-10) to human Ia-like antigen (OE-anti-Ia) is a rapid and convenient method for detection and isolation of Ia-like antigen-bearing cells. Various cell lines and peripheral blood lymphocytes were tested by DRA. Cell lines expressing Ia-like antigen formed rosettes, while cell lines lacking Ia-like antigen did not. In a highly purified T cell fraction, a small number of T cells formed rosettes; but in a Con A-stimulated T cell fraction, about 40% of the cells formed rosettes. The majority of purified B cells and monocytes formed rosettes. Rosette-forming and non-rosette-forming mononuclear cells could be separated by centrifugation on Ficoll-Urografin. Rosette-forming cells reacted positively by indirect immunofluorescence (IIF), while non-rosette-forming cells reacted negatively. The results obtained by DRA were consistent with those obtained by IIF with OKIa*1 and AF-10. DRA is thus suitable not only for detection of Ia-like antigen-bearing cells, but also for separation of Ia-like antigen-positive (Ia+) and -negative (Ia-) cells.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002460:Cell Line; D002469:Cell Separation / Q000379:methods*; D005455:Fluorescent Antibody Technique; D000949:Histocompatibility Antigens Class II / Q000276:immunology*; D006801:Humans; D007962:Leukocytes / Q000276:immunology*; D008214:Lymphocytes / Q000145:classification / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D012397:Rosette Formation / Q000379:methods*
D000911:Antibodies, Monoclonal; D000949:Histocompatibility Antigens Class II