Commercially available antibodies against human and murine histamine H₄-receptor lack specificity

Naunyn Schmiedebergs Arch Pharmacol. 2012 Feb;385(2):125-35. doi: 10.1007/s00210-011-0700-4. Epub 2011 Nov 10.


Antibodies are important tools to detect expression and localization of proteins within the living cell. However, for a series of commercially available antibodies which are supposed to recognize G-protein-coupled receptors (GPCR), lack of specificity has been described. In recent publications, antisera against the histamine H₄-receptor (H₄R), which is a member of the GPCR family, have been used to demonstrate receptor expression. However, a comprehensive characterization of these antisera has not been performed yet. Therefore, the purpose of our study was to evaluate the specificity of three commercially available H₄R antibodies. Sf9 insect cells and HEK293 cells expressing recombinant murine and human H₄R, spleen cells obtained from H₄⁻/⁻ and from wild-type mice, and human CD20⁺ and CD20⁻ peripheral blood cells were analyzed by flow cytometry and Western blot using three commercially available H₄R antibodies. Our results show that all tested H₄R antibodies bind to virtually all cells, independently of the expression of H₄R, thus in an unspecific fashion. Also in Western blot, the H₄R antibodies do not bind to the specified protein. Our data underscore the importance of stringent evaluation of antibodies using valid controls, such as cells of H₄R⁻/⁻ mice, to show true receptor expression and antigen specificity. Improved validation of commercially available antibodies prior to release to the market would avoid time-consuming and expensive validation assays by the user.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies / immunology*
  • Antibody Specificity
  • Baculoviridae / genetics
  • Cell Line
  • HEK293 Cells
  • Humans
  • Leukocytes, Mononuclear / immunology
  • Mice
  • Mice, Inbred BALB C
  • Mice, Knockout
  • Receptors, G-Protein-Coupled / deficiency
  • Receptors, G-Protein-Coupled / genetics
  • Receptors, G-Protein-Coupled / immunology*
  • Receptors, Histamine / deficiency
  • Receptors, Histamine / genetics
  • Receptors, Histamine / immunology*
  • Receptors, Histamine H4
  • Spodoptera
  • Transfection


  • Antibodies
  • HRH4 protein, human
  • Hrh4 protein, mouse
  • Receptors, G-Protein-Coupled
  • Receptors, Histamine
  • Receptors, Histamine H4