IMMUNOHISTOCHEMICAL DETECTION OF THE HUMAN MAJOR VAULT PROTEIN LRP WITH 2 MONOCLONAL-ANTIBODIES IN FORMALIN-FIXED, PARAFFIN-EMBEDDED TISSUES

Multidrug resistant cancer cells frequently overexpress the 110-kd lun g resistance-related protein (LRP) as detected with the monoclonal ant ibody (MAb) LRP-56. Recently, we identified LRP as the major vault pro tein (MVP), which is the major constituent of vaults, multisubunit cel lular organelles. Clinically, LRP/MVP expression in cancer at time of diagnosis provided a strong and independent prognostic factor for resp onse to chemotherapy and outcome in different tumor types, notably acu te myeloid leukemia and ovarian cancer. To facilitate additional immun ohistopathological studies, we have optimized LRP/MVP detection in par affin-embedded tissues using two monoclonal antibodies, LRP-56 and LMR -5, Blocking experiments showed that LRP-56 and LMR-5 MAbs detect diff erent epitopes of LRP/MVP, Immunohistochemical studies with both MAbs in a panel of human multidrug resistant tumor cell lines, normal tissu es, and colorectal tumors showed that LRP/MVP expression can be reliab ly detected after formalin-fixation and paraffin-embedding using overn ight incubation at 4 degrees C with the primary MAbs at 5- to 10-fold higher concentrations (ie, 1 to 10 mu g/ml) as currently used with fro zen sections, Both streptavidin-biotin complex and alkaline phosphatas e-anti-alkaline phosphatase techniques could be successfully used for signal-amplification, Staining quality did not benefit from antigen-re trieval pretreatments. The optimized staining methodology facilitates studies in archival material on the putative role of LRP/MVP in clinic al drug resistance.