Detection of rare human breast cancer cells. Comparison of an immunomagnetic separation method with immunocytochemistry and RT-PCR.
BACKGROUND
The detection of occult carcinoma cells in patients with breast cancer may aid determination of prognosis and the development of new therapeutic approaches. In this study, we report a new method to detect rare human breast cancer cells, which combines an immunomagnetic separation (IMS) procedure with cytokeratin 19 (CK 19) immunostaining.


METHODS
Four monoclonal antibodies (MAb) previously characterized against cell surface antigens (1BE12, ED8, 7B10 and 83D4), were evaluated for IMS optimization. Immunoseparated epithelial cells were identified using a MAb against CK 19. We compared the IMS procedure with the immunocytochemistry (ICC) and the RT-PCR for CK 19 on an "in vitro" experimental model.


RESULTS
The best results in IMS procedures were obtained using MAbs 1BE12 (directed against Lewis y antigen) and ED8 (directed against MUC 1). In reconstitution experiments, using several ratios of T47D cells mixed with peripheral-blood mononuclear (PBMN) cells, the IMS procedure reliably detects one mammary carcinoma cell in 5 x 10(5) PBMN cells, whereas the ICC detects up to one T47D cell per 10(5) PBMN cells. The best sensitivity was observed with the RT-PCR (up to one T47D cell per 10(6) PBMN cells). We found the same high specificity with the three methods evaluated.


CONCLUSIONS
The IMS procedure using MAbs 1BE12 or ED8 associated with CK 19 immunostaining is a specific, sensitive, and feasible method for the detection of rare human breast cancer cells. This method proved to be better than the ICC staining but its sensitivity was lower than that of RT-PCR for CK 19.
Berois
N
N
Depto. de Bioquímica, Facultad de Medicina, Montevideo, Uruguay.
Varangot
M
M
Osinaga
E
E
Babino
A
A
Caignault
L
L
Musé
I
I
Roseto
A
A
D000911:Antibodies, Monoclonal; D001943:Breast Neoplasms / Q000175:diagnosis* / Q000473:pathology; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D007150:Immunohistochemistry; D018189:Immunomagnetic Separation; D007633:Keratins / Q000235:genetics / Q000378:metabolism; D018365:Neoplasm, Residual / Q000175:diagnosis*; D016133:Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D007633:Keratins