Application of the avidin-biotin-complex method for the light microscopic analysis of lymphocyte subsets with monoclonal antibodies on air-dried smears.
A study was undertaken of the application of the avidin-biotin-peroxidase complex (ABC) method to the monoclonal antibody MAbs staining of mononuclear cells in hematologic and cytodiagnostic materials. Satisfactory cell morphology and immunoreactivity of surface antigens were observed when the slides were fixed in 80% acetone in phosphate-buffered saline or in 60% acetone in 0.03 M citric acid buffer solution (pH 5.4). Unstained air-dried preparations could be preserved for two weeks at room temperature in a desiccator and for one year at -70 degrees C after fixation. An excellent immunoreaction, even with a weak surface antigen, was observed by inhibition of endogenous peroxidase after the secondary antibody reaction; reactions of weak antigens tended to be obscured when the inhibition was performed before the first antibody reaction. Use of the Giemsa stain as a counterstain made it possible to readily observe the cell morphology; therefore, white blood cell analysis could be performed simultaneously when peripheral blood smears were studied. The positive rate of immunoreaction by an immunofluorescent method was well correlated with that obtained by the ABC method. The ABC method proved to be an excellent immunocytochemical technique for detecting cell surface antigens with high sensitivity and specificity; furthermore, it is useful for cell morphology studies and yields permanent preparations.
Aratake
Y
Y
Central Laboratory for Clinical Investigation, Miyazaki Medical College Hospital, Japan.
Tamura
K
K
Kotani
T
T
Ohtaki
S
S
D000911:Antibodies, Monoclonal; D000941:Antigens / Q000032:analysis; D003581:Cytodiagnosis; D005455:Fluorescent Antibody Technique; D006801:Humans; D007136:Immunoglobulins / Q000032:analysis; D007202:Indicators and Reagents; D007938:Leukemia / Q000276:immunology*; D007945:Leukemia, Lymphoid / Q000276:immunology; D008214:Lymphocytes / Q000145:classification* / Q000166:cytology / Q000276:immunology; D008223:Lymphoma / Q000276:immunology / Q000473:pathology; D008581:Meningitis / Q000209:etiology / Q000276:immunology / Q000473:pathology; D009107:Mumps / Q000276:immunology / Q000473:pathology; D012016:Reference Values
D000911:Antibodies, Monoclonal; D000941:Antigens; D007136:Immunoglobulins; D007202:Indicators and Reagents