Monoclonal antibodies to denatured human ACE (CD 143), broad species specificity, reactivity on paraffin sections, and detection of subtle conformational changes in the C-terminal domain of ACE.
Two new mouse monoclonal antibodies (mAbs) were generated to denatured human angiotensin-converting enzyme (ACE, CD143). The clones 2E2 and 3C5, each of the IgG1 kappa chain isotype, detect ACE with high sensitivity, respectively, at 20 ng and 2 ng of protein per lane in Western blotting. They both recognize different epitopes on the C-domain of ACE located between amino acid residues 740 and 992. In formalin-fixed and paraffin-embedded human tissues, immunohistochemistry revealed all known expression sites of ACE, e.g. the epithelial brush borders of proximal kidney tubules, epithelial cells of epididymis, endothelial cells, activated macrophages as well as germ cells during spermatogenesis. In contrast to other mAbs to denatured human ACE, mAbs 2E2 and 3C5 demonstrate cross-reactivity with a broad spectrum of animal species such as monkey, rat, rabbit, cattle, dog, cat, and guinea pig. In addition, mAb 2E2 recognized mouse ACE in Western blotting and on paraffin sections. Our findings suggest that mAbs 2E2 and 3C5 are useful for identifying even subtle changes in ACE conformation resulting from denaturation. These mAbs are also sensitive tools for the detection of minimal amounts of ACE in biological fluids and tissues using proteomics approaches. Their reactivity in routinely processed tissues of various species may prove useful for correlation of ACE expression in animal models to human diseases.
Balyasnikova
I V
IV
Department of Anesthesiology, University of Illinois at Chicago, IL 60612, USA.
Metzger
R
R
Franke
F E
FE
Danilov
S M
SM
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015153:Blotting, Western; D002415:Cats; D002417:Cattle; D003429:Cross Reactions; D004285:Dogs; D006168:Guinea Pigs; D006801:Humans; D006825:Hybridomas; D007150:Immunohistochemistry; D051379:Mice; D008969:Molecular Sequence Data; D016612:Paraffin Embedding; D007703:Peptidyl-Dipeptidase A / Q000737:chemistry / Q000276:immunology*; D010969:Plastics / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D011489:Protein Denaturation; D017434:Protein Structure, Tertiary / Q000502:physiology; D011817:Rabbits; D051381:Rats; D016415:Sequence Alignment; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D010969:Plastics; D007703:Peptidyl-Dipeptidase A