[Parameters of specific antibody interaction with P-gp in T-cell leukemia Jurkat cells].
Special features of Pgp expression evaluation by flow cytometry were investigated. Indexes of interaction of FITC-conjugated Becton Dickinson Pharmingen monoclonal antibodies to external Pgp epitope (clone 17F9) were analyzed depending on the cell concentration (400000 to 3000000 cells/ml) and the specific antibody concentration (5, 10 and 20 microl of the market product solution per 300 microl of the cell suspension).


RESULTS
1. Optimal condition of incubation with the antibodies was revealed--after the cell fixation in 4% formaldehyde. 2. Character of the increase of the cell fluorescence average intensity in the suspension totally according to the concentration of the Pgp-specific antibodies did not depend on the number of the cells. 3. Both the absolute value of the average intensity of the cell specific fluorescence as well as cell number out of the isotypic control fluorescence region depended on the ratio of the cell number to monoclonal antibody concentration.


CONCLUSIONS
1. It was shown that Pgp was practically expressed in all Jurkat cells. 2. By the Pgp expression level, the Jurkat cell culture was sufficiently homogeneous and stable in various passages. 3. Jurkat cells could be used as test culture in estimation of the market antibody activity. 4. For immunofluorescent assay of the Pgp expression in human tumor biopsy specimens, it is necessary to use not less than three concentrations of the specific antibodies, not less than three concentrations of the cells in the suspension as well as concurrent assay of the cell culture characterized previously. In particular, for investigated Pgp monoclonal antibodies, it is possible to use Jurkat cell culture. It allows revealing not only the fact of the Pgp expression but the level of the expression as well, i.e. to estimate severity of multidrug resistance phenotype.
Bogush
T A
TA
Dudko
E A
EA
Bogush
E A
EA
Baryshnikov
A Iu
AIu
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis* / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity / Q000276:immunology*; D014408:Biomarkers, Tumor / Q000032:analysis* / Q000276:immunology; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D000939:Epitopes / Q000276:immunology; D005434:Flow Cytometry / Q000379:methods*; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D006801:Humans; D019169:Jurkat Cells; D012680:Sensitivity and Specificity
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000911:Antibodies, Monoclonal; D014408:Biomarkers, Tumor; D000939:Epitopes; D005456:Fluorescent Dyes; D016650:Fluorescein-5-isothiocyanate