Characterization and Applications of a Newly Developed Rabbit Monoclonal Antibody to Cytokeratin 7 (CK7) for Immunohistochemistry

Abstract

In this paper, we describe the development of a novel rabbit monoclonal cytokeratin 7 (CK7) antibody. CK7 is widely used in immunohistochemical tests that are commonly performed in histopathology laboratories. Rabbits were immunized with a peptide corresponding to C-terminus of CK7. Recombinant monoclonal antibodies were developed using proprietary technology. Antibodies were screened by enzyme-linked immunosorbent assay against CK7 peptide, and the specificity of the antibody was verified by Western blotting on Hela cell lysates and by immunohistochemical staining on formalin-fixed paraffin-embedded tissue sections. One clone, designated as BC1 was selected and tested on a variety of human tissues. Its reactivity was tested and compared with a well-characterized mouse monoclonal CK7 antibody OV-TL 12/30 on lung, breast, ovarian, and colon cancers, and on a variety of normal tissues. The staining patterns and specificity were concordant with mouse monoclonal OV-TL 12/30. The BC1 clone, however, showed superior staining to OV-TL 12/30 in terms of intensity and staining of debris. CK7 combined with CK20 or CDX2 and TTF1 is a common panel used in histopathology laboratories in discrimination between primary adenocarcinomas and metastatic lung adenocarcinomas from colonic origin. We then developed a 4-step double stain (multiplex) assay by combining CDX-2 with CK7 in an antibody cocktail and tested the efficacy on lung, breast, and colon cancers. Results showed that this novel antibody can be used as a single component and/or in combination with CDX2 and TTF1 in applications for diagnostic pathology.