Characterization of a new antibody raised against the NH2 terminus of P-glycoprotein

Prema S. Rao, Rajgopal Govindarajan, Kavita B. Mallya, William Walter West, U. Subrahmanyeswara Rao

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Purpose: Cancers exposed to chemotherapy develop multidrug resistance, a major cause for chemotherapy failure. One mechanism of multidrug resistance development is due to overexpression of P-glycoprotein (Pgp) in these cancer cells. Thus, a prechemotherapy evaluation of Pgp in cancer cells aids in the design of alternative regimens that can circumvent such failure. As few Pgp-specific antibodies are available in detecting low levels of Pgp, there is a need for preparing an antibody that allows the detection of Pgp by various immunologic methods. Experimental Design: We selected the amino acid stretch 11 to 34 in the cytoplasmically located NH2 terminus of Pgp as antigen, which was chemically synthesized and used to raise an antibody in a rabbit, termed NH211 antibody. We compared the properties of NH211 antibody with that of the well-characterized Pgp-specific antibody, C219, by Western blotting, immunoprecipitation, immunocytochemistry, and immunohistochemistry. Results: Immunoblotting analysis suggested that NH 211 antibody efficiently interacts with both recombinant and constitutively expressed Pgp in cancerous and noncancerous human cells. Immunoprecipitation reactions indicated that the NH211 antibody selectively immunoprecipitates Pgp. Immunocytochemical analyses indicated that the NH211 antibody detects Pgp in drug-resistant breast cancer cells as well as in human prostate and breast adenocarcinoma tissue sections. Conclusion: As the NH211 antibody detects Pgp present in cells and tissues, we conclude that the amino acid sequence to which this antibody was raised is highly antigenic and the antibody is useful in the detection of Pgp by a variety of immunologic methods.

Original languageEnglish (US)
Pages (from-to)5833-5839
Number of pages7
JournalClinical Cancer Research
Volume11
Issue number16
DOIs
StatePublished - Aug 15 2005

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P-Glycoprotein
Antibodies
Multiple Drug Resistance
Immunoprecipitation
Immunohistochemistry
Drug Therapy
Neoplasms
Immunoblotting
Prostate
Amino Acid Sequence
Adenocarcinoma
Breast
Research Design
Western Blotting
Breast Neoplasms
Rabbits
Antigens
Amino Acids

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

Rao, P. S., Govindarajan, R., Mallya, K. B., West, W. W., & Rao, U. S. (2005). Characterization of a new antibody raised against the NH2 terminus of P-glycoprotein. Clinical Cancer Research, 11(16), 5833-5839. https://doi.org/10.1158/1078-0432.CCR-04-2182

Characterization of a new antibody raised against the NH2 terminus of P-glycoprotein. / Rao, Prema S.; Govindarajan, Rajgopal; Mallya, Kavita B.; West, William Walter; Rao, U. Subrahmanyeswara.

In: Clinical Cancer Research, Vol. 11, No. 16, 15.08.2005, p. 5833-5839.

Research output: Contribution to journalArticle

Rao, Prema S. ; Govindarajan, Rajgopal ; Mallya, Kavita B. ; West, William Walter ; Rao, U. Subrahmanyeswara. / Characterization of a new antibody raised against the NH2 terminus of P-glycoprotein. In: Clinical Cancer Research. 2005 ; Vol. 11, No. 16. pp. 5833-5839.
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