title: Multiparameter flow cytometric measurement of epidermal growth factor receptor and c-erbB-2 oncoprotein in cultured cells and in fresh and preserved solid tumor cells.
abstract: We developed a multiparameter flow cytometric technique for the simultaneous measurement of cellular DNA content and c-erbB-2 or epidermal growth factor receptor (EGFR) expression. The method provides a high resolution of DNA content and well preserved c-erbB-2 and EGFR immunostaining under saturated antibody conditions, allowing good control for background fluorescence and satisfactory cell morphology. Four different protocols for the short-term preservation of cells used for multiparameter flow cytometric assay of EGFR and c-erbB-2 were assessed in cell suspensions prepared by mechanical disaggregation in 10 gynecologic tumors. The protocols at 4 degrees C were: storage in 50% methanol, and storage in buffer after formalin fixation. Tissues were also cryopreserved as cell suspensions and tissue blocks. When the oncoprotein expression and DNA histograms were compared with those in fresh suspensions, cryopreservation was found to be the best method: oncoprotein expression was well preserved and there was a good correlation between oncoprotein expression and the quality of the DNA histograms. The currently developed methods for cell preservation make the technique generally available for clinical cancer studies.
authors.affiliation: Department of Gynaecologic Oncology and Histopathology, St Bartholomew's Hospital, London, UKDepartment of Obstetrics and Gynaecology, Antwerp University Hospital, Edegem, BelgiumMedical Research Council, Clinical Oncology Unit, Cambridge, UK.
authors.forename: P.A.
authors.initials: PA
authors.lastname: van Dam
authors.forename: D.G.
authors.initials: DG
authors.lastname: Lowe
authors.forename: J.V.
authors.initials: JV
authors.lastname: Watson
authors.forename: J.H.
authors.initials: JH
authors.lastname: Shepherd