A New Rabbit Monoclonal Antibody Against PAX2
Aihua Li, Nenghui Jiang, Hongyang Pan, Asish Nand, Juliet Gardiner, Zhiling Fang, Taiying Chen. Epitomics, Inc., Burlingame, CA
Background: The PAX2 gene belongs to the paired box gene family that plays a critical role in the formation of the kidney, eye, ear, central nervous system and urogenital tract during embryonic development. After birth, PAX2 is involved in protecting against cell death during periods of cellular stress. PAX2 has been used as a marker for the identification of renal cell carcinoma and ovary carcinoma by immunohistochemistry (IHC). However, current commercially available PAX2 antibodies have been shown to have cross reactivity in lymphomas and other tumors. The superior sensitivity and specificity of rabbit monoclonal antibodies (RabMAbs) in the IHC detection of antigens in formalin-fixed paraffin-embedded (FFPE) tissue, as compared to mouse monoclonal and rabbit polyclonal antibodies, has been attributed to several advantages of the rabbit immune system in response to antigen immunization. Here we report the development of a PAX2 antibody that specifically labels target cells, by utilizing the RabMAb technology.
Design: Rabbits were immunized with recombinant PAX2 protein. Sera collected from immunized rabbits were screened by ELISA, western blot (WB) and IHC. After fusion, antibody from final hybridoma cell line was further characterized by extensive IHC testing using FFPE human normal, tumor and PAX2 positive tumor tissues.
Results: A 45 KDa PAX2 protein was confirmed by WB. IHC analysis on human tissue microarray (TMA) comprising twenty-four types of adult and ten types of fetal normal tissues showed that this PAX2 antibody reacts with epithelial cells in kidney, fallopian tube, ovary, epididymis, and vesicula seminalis in the nuclei. No staining was observed in other types of cells. In a urinary tumor TMA, PAX2 was positive in 10 out of 15 kidney clear cell tumors, 2 out of 2 Wilms' tumors, but negative in 3 of out of 3 transitional cell carcinomas. In tumor TMA comprising twenty-three types of other tumor tissues, no PAX2 positive staining was observed. In contrast to a commercial rabbit polyclonal anti-PAX2 antibody, this PAX2 antibody shows no immunoreactivity in normal and neoplastic lymphocytes.
Conclusions: The RabMAb PAX2, clone EP235 specifically labels epithelial cells in the kidney and genital tract and tumors with kidney origin. Compared to currently available PAX2 antibody, no immunoreactivity in lymphoid cells was observed. This PAX2 antibody may be a better tool in identifying cancers with kidney origin on FFPE tissues.
Category: Genitourinary (including renal tumors)
Wednesday, March 6, 2013 1:00 PM
Poster Session VI # 177, Wednesday Afternoon