Measurement of P-glycoprotein expression in multidrug-resistant human neuroblastoma cell lines using self-competitive binding assay Journal Article
| Authors: | Levchenko, A.; Mehta, B. M.; Spengler, B. A.; Narkar, A. A.; Fonti, R.; Biedler, J. L.; Tsuruo, T.; Larson, S. M. |
| Article Title: | Measurement of P-glycoprotein expression in multidrug-resistant human neuroblastoma cell lines using self-competitive binding assay |
| Abstract: | To date, all reported measurements of multidrug resistance have been semiquantitative. The purpose of the present study is to establish and validate the self-competitive binding assay technique utilizing monoclonal antibody for quantitative estimation of multidrug resistance in tumor cells. This technique is used for P-glycoprotein concentration measurement in BE(2)- C human neuroblastoma cell line and its sublines with primary resistance to colchicine and actinomycin D. Monoclonal antibody MRK-16 was used in this study. It was labeled with iodine-125 (125I) to trace the concentration of antibody-antigen complexes. The binding data were obtained by varying the concentration of the unlabeled antibody. The results were fitted to a model equation to estimate the number of binding sites and antibody-antigen dissociation constant. The P-glycoprotein concentration was significantly higher in the resistant sublines than in the sensitive line. The highest levels were achieved in actinomycin D-resistant cells: 2.1 x 106 binding sites/cell versus 5.4 x 104 binding sites/cell in the sensitive cells. The consistency of the results was verified by repeating the study three times for each cell line. The binding assay results were confirmed by Western blot experiments performed on the same cell lines. |
| Keywords: | human cell; doxorubicin; gene expression; vincristine; monoclonal antibody; iodine 125; neuroblastoma cell; dactinomycin; binding site; protein determination; multidrug resistance; binding competition; colchicine; dissociation constant; glycoprotein p; antigen antibody complex; quantitative diagnosis; human; priority journal; article |
| Journal Title: | Analytical Biochemistry |
| Volume: | 236 |
| Issue: | 2 |
| ISSN: | 0003-2697 |
| Publisher: | Academic Press, Elsevier Inc |
| Date Published: | 1996-05-01 |
| Start Page: | 338 |
| End Page: | 343 |
| Language: | English |
| DOI: | 10.1006/abio.1996.0176 |
| PUBMED: | 8660514 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | Article -- Export Date: 22 November 2017 -- Source: Scopus |
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