Flow cytometric evaluation of multidrug resistance proteins

Abstract

There are several ways to detect proteins on cells. One quite frequently used method is flow cytometry. This method needs fluorescently labeled antibodies that can attach selectively to the protein to be investigated for flow cytometric detection. Flow cytometry scans individual cells, virtually without their surrounding liquid, and can scan many cells in a very short time. Because of this advantage of flow cytometry, it was adapted to investigate transport proteins on normal and cancerous human cells and cell lines. These transport proteins play important roles in human metabolism. Absorption in the intestine, excretion at the kidney, protection of the CNS compartment and the fetus from xenobiotics, and other vital functions depend on these transporters. However, several transporters are overexpressed in cancer cells. These overexpressed transporters pump out anticancer drugs from the cells and prevent their curative effects. The detection and quantitation of these types of transporters in cancer cells is important for this reason. Here, we review literature on flow cytometric detection of the three most studied transporters: P-glycoprotein, multidrug resistance-associated proteins, and breast cancer resistance protein.

Fig. 7.1.

Primary structure of P-glycoprotein as positioned in cell membrane, with three antibodies (MRK16, UIC-2, and 17F9) positioned at the known binding epitopes (Adapted from Ambudkar et al. (2003) P-glycoprotein: from genomics to mechanism. Oncogene 22:7468–7485, with permission of the author).

Fig. 7.2.

Differences in histogram intensity after binding anti-mouse FITC, MRK16-FITC, or MRK-16 + anti-mouse FITC to P-glycoprotein-expressing NIH3T3MDR cells (From Aszalos and Weaver (1998) Estimation of drug resistance by flow cytometry. In: Jaroszeski and Heller (eds) Flow cytometry protocols. Humana, Totowa, NJ, pp 117–122, Figure 2, p. 121, with kind permission of Springer Science).

Fig. 7.3.

Influence of inhibitors on the side population of A549 cells stained with Hoechst 33342 dye (From Scharenberg et al. (2002) The ABCG2 transporter is an efficient Hoechst 33342 efflux pump and is preferentially expressed by immature human hematopoietic progenitors. Blood 99:507–512, used by permission).