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Flow Cytometric Analysis of Glucocorticoid Receptor Using Monoclonal Antibody and Fluoresceinated Ligand Probes¹

Department of Hematology, M. D. Anderson Hospital and Tumor Institute, Houston, Texas 77030 [D. M., N. T. V., B. B.]; Department of Human Biological Chemistry and Genetics, The University of Texas Medical Branch at Galveston, Galveston, Texas 77550 [B. G., E. B. T.]; and Clinical Chemistry Laboratory, Kobe Women's College of Pharmacology, Kobe, Japan [Y. K., F. W.]

Conditions were established for single cell analysis of glucocorticoid receptor (GR) content by flow cytometry using several clones of a human leukemic cell line (CCRF-CEM). These included CEM-7A, 7R, C1, and ICR 27 Tk.3 cells which were examined both by standard [³H]dexamethasone radiometric binding and by two independent flow cytometry assays. The latter involved either mouse monoclonal antibody against GR (GR-MoAb) or fluoresceinated cortisol ligand probes. For CEM-7A, 7R, and C1 cells, there was a correlation between GR-MoAb and radiometrically defined GR values. However, clone ICR-27 Tk.3 with low [³H]dexamethasone binding exhibited the highest GR-MoAb fluorescence. The fluoresceinated cortisol assay correlated with dexamethasone binding values in all four clones. Thus, GR-MoAb identifies the total immunologically reactive GR present, while the fluoresceinated cortisol assay quantifies only the functionally intact GR in terms of its initial binding. Their combined use may reveal the cellular heterogeneity of GR expression and function also in human tumor samples, to which they have been successfully applied. When coupled with DNA counterstaining, GR expression can be related directly to frequently DNA-aneuploid tumor cells and cell cycle distribution.

¹ Supported in part by Grants CA 28771 and CA 16672 from the NIH, Bethesda, MD. This work was also done partly in conjunction with the Walls Medical Research Foundation.

² To whom requests for reprints should be addressed, at Department of Hematology, M. D. Anderson Hospital, 1515 Holcombe Blvd., Houston, TX 77030.

Received 9/29/87. Revised 4/29/88. Revised 10/13/88. Accepted 10/29/88.