VERSATILITY OF STAPHYLOCOCCUS-AUREUS PROTEIN-A IN IMMUNOCYTOCHEMISTRY - USE IN UNLABELED ANTIBODY ENZYME-SYSTEM AND FLUORESCENT METHODS

被引:32
作者
NOTANI, GW [1 ]
PARSONS, JA [1 ]
ERLANDSEN, SL [1 ]
机构
[1] UNIV MINNESOTA,SCH MED,DEPT ANAT,MINNEAPOLIS,MN 55455
关键词
D O I
10.1177/27.11.390040
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
A modification of the Sternberger PAP technique for immunocytochemical staining utilizing the binding of protein A (PA) to the Fc component of IgG in place of the specific anti-IgG bridge step has been tested. We have found PA to be useful in the detection of a variety of antigens including cell surface and cytoplasmic antigens of peripheral blood leucocytes and spleen cells, tissue sections of pituitary, pancreas and intestine and viral antigens of tissue culture cells. Also, PA can be used for immunoenzyme counterstaining of fluorescent preparations and for antibody titrations. Pa can be used to detect primary antibody immune complexes in a wide variety of species and thus eliminates the need for individual anti-IgG antisera and corresponding PAP immune complexes. PA bound to the tissue antigens through IgG immune complexes showed high affinity for rabbit and guinea pig PAP and lower affinity for rat and goat PAP. The high affinity of PA for rabbit IgG could also be demonstrated as all the Fc receptors of the tissue bound PA could be saturated with rabbit IgG. For immunofluorescence of cellular antigens, fluorescein-conjugated PA could be used directly or PA could be used as a bridge between primary antibody immune complex and a fluorescent conjugate of IgG. In both cases, the resulting immunofluorescent staining could be made permanent by immunoenzyme staining with PAP or with PA and PAP, respectively. Antibody titrations were compared using the binding of guinea pig anti-growth hormone (GH) antiserum over a 100- t0 50,000-fold range of dilutions determined by PA-PAP immunocytochemistry and by RIA. The two methods yielded similar titration curves and sensitivity of detection. The time course kinetics from 1 to 15 minutes for the binding of primary antibody detected in the immunoenzyme system could be produced with a 2500-fold dilution of primary antibody. The rapid course of immunostaining coupled with the versatility imparted by the Fc binding of the PA reagents suggests that PA immunoenzyme staining should be suitable for a variety of antigens and for clinical evaluation of cell surface antigens such as immunoglobulins.
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页码:1438 / 1444
页数:7
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