MULTIPARAMETER FLOW CYTOMETRIC MEASUREMENT OF EPIDERMAL GROWTH-FACTORRECEPTOR AND C-ERBB-2 ONCOPROTEIN IN CULTURED-CELLS AND IN FRESH AND PRESERVED SOLID TUMOR-CELLS

We developed a multiparameter flow cytometric technique for the simult aneous measurement of cellular DNA content and c-erbB-2 or epidermal g rowth factor receptor (EGFR) expression. The method provides a high re solution of DNA content and well preserved c-erbB-2 and EGFR immunosta ining under saturated antibody conditions, allowing good control for b ackground fluorescence and satisfactory cell morphology. Four differen t protocols for the short-term preservation of cells used for multipar ameter flow cytometric assay of EGFR and c-erbB-2 were assessed in cel l suspensions prepared by mechanical disaggregation in 10 gynecologic tumors, The protocols at 4 degrees C were: storage in 50% methanol, an d storage in buffer after formalin fixation. Tissues were also cryopre served as cell suspensions and tissue blocks. When the oncoprotein exp ression and DNA histograms were compared with those in fresh suspensio ns, cryopreservation was found to be the best method: oncoprotein expr ession was well preserved and there was a good correlation between onc oprotein expression and the quality of the DNA histograms. The current ly developed methods for cell preservation make the technique generall y available for clinical cancer studies.