Quantitative cytofluorimetric determination of cell membrane-associated large tumor antigen on SV40-transformed cells.
The aim of this study was to quantitate the number of cell membrane-located SV40 large tumor antigen (large T) molecules of SV40-transformed cell lines by cytofluorimetric analysis. Five different SV40-transformed cell lines were labelled by either a biotin- or a fluorescein-conjugated monoclonal antibody, PAb1605, which is specific for the large T carboxyterminus. The conjugated-antibody fluorescence signals of the stained large T molecules of transformed cells were measured via cytofluorimetry. Comparison of the fluorescence signals of calibrated beads bearing a known number of fluorescein molecules to the signals of conjugated PAb1605 antibodies bound on microbeads to a defined number of IgG binding sites made it possible to determine the number of antibody-accessible large T molecules per SV40-transformed cell. The numbers (x10(-4)) found per cell were 1.0 (ELONA, hamster), 3.0 (VLM, mouse), 3.5 (mKSA, mouse), 11 (C57SV, mouse), and 5.5 (SV80, human), respectively. Thus, the technique described allows a precise quantitation of surface-exposed, antibody-accessible viral antigen expression.
Hess
R D
RD
Abteilung Virologie, Albert-Ludwigs-Universität, Freiburg, Germany.
Kuther
M
M
Haessler
C
C
Paetzold
S
S
Braun
D G
DG
Brandner
G
G
D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000032:analysis* / Q000276:immunology; D002461:Cell Line, Transformed; D002462:Cell Membrane / Q000276:immunology*; D002467:Cell Nucleus / Q000276:immunology; D006224:Cricetinae; D005434:Flow Cytometry / Q000379:methods*; D005455:Fluorescent Antibody Technique; D006801:Humans; D051379:Mice
D000952:Antigens, Polyomavirus Transforming