A new immunohistochemical methodology for the specific detection of MDR1-P-glycoprotein in human tissues based on phage-displayed peptides mimicking the MM4.17 epitope.
It is not rare that controversial indications about the presence or the expression level of multidrug-resistant (MDR) proteins come out from different laboratories upon examination of identical tumor specimens. Distinct aspects, including the use of weakly discriminating monoclonal antibodies (MAbs) and/or unsuitable techniques and procedures, contribute in generating differences in the MDR phenotype evaluation of cancer cells. In this regard we describe here an innovative immunohistochemical approach for the determination of P-glycoprotein expression in cells and tissues. The method is based on the ability of phage-displayed peptides to mimic antibody epitopes. For this purpose we utilized the phage clone #55, which was affinity-purified from a phage-displayed random-peptide library using the MAb MM4.17 (specific for MDR1-P-glycoprotein) as previously described. This clone has been chosen since it clearly and undoubtedly reacts with its cognate MAb, as was determined by ELISA and dot blot tests. Inhibition of the MAb MM4.17 binding to MDR1-P-glycoprotein-expressing cells could be performed by adding a calibrated concentration of phage clone #55 particles, which mimic MDR1-P-glycoprotein antigen. This methodology can eliminate misleading interpretations concerning the presence and expression level of MDR1-P-glycoprotein and might well contribute in routine clinical determinations of MDR in tumor specimens, thus contributing to our understanding of the basis of the mechanisms of tumor cell resistance to drugs.
Poloni
F
F
Laboratorio di Immunologia, Istituto Superiore di Sanità, Roma, Italy.
Castagna
M
M
Felici
F
F
Cianfriglia
M
M
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D000911:Antibodies, Monoclonal / Q000737:chemistry; D000918:Antibody Specificity; D001435:Bacteriophages / Q000737:chemistry / Q000276:immunology*; D001667:Binding, Competitive / Q000276:immunology; D018432:Drug Resistance, Multiple / Q000276:immunology; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry / Q000379:methods*; D007668:Kidney / Q000166:cytology / Q000276:immunology; D008099:Liver / Q000166:cytology / Q000276:immunology; D009928:Organ Specificity / Q000276:immunology; D019151:Peptide Library; D013194:Staining and Labeling
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000911:Antibodies, Monoclonal; D000939:Epitopes; D019151:Peptide Library