Development and characterization of rat monoclonal antibodies to denatured mouse angiotensin-converting enzyme.
Four new rat monoclonal antibodies, generated to denatured mouse somatic angiotensin-converting enzyme (ACE, CD143), detect mouse ACE with high sensitivity in Western blotting. Epitope mapping for the monoclonal antibodies--B12, 4G6 and 5C4--was also performed. Two monoclonal antibodies--B12 and 5C4--are directed to various epitopes on the N-domain--i.e., they recognized only the somatic isoform of mouse ACE. The monoclonal antibody H7 recognized an epitope on the C-domain of mouse ACE. The monoclonal antibody 4G6 was directed to a sequence on the N-domain of mouse ACE, which is homologous to a region of the C-domain and, as a result, also recognizes mouse testicular ACE (tACE) by means of Western blotting. In paraffin-embedded mouse tissues, all monoclonal antibodies detected all known expression sites of somatic ACE (sACE), e.g., the epithelial cells of the kidney proximal tubules, intestine and epididymis, and heterogeneously in endothelial cells. The monoclonal antibodies 4G6 and H7 additionally stained mouse tACE in spermatozoa and in mature spermatids. The monoclonal antibody 4G6 also demonstrated cross-reactivity with sACE from a broad spectrum of animal species, including human, rat, rabbit and bovine. However, this monoclonal antibody did not recognize the testicular isoform of ACE of these species. This set of monoclonal antibodies is useful for identifying even subtle changes in mouse ACE conformation because of denaturation. These monoclonal antibodies are also sensitive tools for the detection of mouse ACE in biological fluids and tissues by using proteomics approaches. Their high reactivity in paraffin-embedded tissues opens up opportunities to study possible changes in the pattern of ACE expression in knockout mouse models and may prove useful for correlating ACE expression in these models with human diseases.
Balyasnikova
I V
IV
Department of Anesthesiology, University of Illinois at Chicago, Chicago, IL 60612, USA.
Metzger
R
R
Sun
Z-L
ZL
Berestetskaya
Y V
YV
Albrecht
R F
RF
Danilov
S M
SM
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015153:Blotting, Western; D002417:Cattle; D003429:Cross Reactions; D018604:Epitope Mapping; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D007703:Peptidyl-Dipeptidase A / Q000032:analysis / Q000737:chemistry / Q000276:immunology*; D011489:Protein Denaturation; D017434:Protein Structure, Tertiary; D011817:Rabbits; D051381:Rats; D014018:Tissue Distribution
D000911:Antibodies, Monoclonal; D007703:Peptidyl-Dipeptidase A