New monoclonal antibodies against bilitranslocase as a diagnostic tool in determining the progress of clear cell renal cell carcinoma

Abstract

Background: Monoclonal antibodies (mAbs) are an important tool in diagnostics and research, especially when we are dealing with a protein marker of unknown primary structure as in the case of bilitranslocase (BTL). BTL is also expressed on kidney cells, where it acts as an organic anion transporter. We have shown earlier that there are differences in bilitranslocase expression in normal kidney cells versus early grade kidney cancer.Methods: We developed monoclonal antibodies against extra- and intra-cellular domains of bilitranslocase protein model. To also gain a deeper insight in bilitranslocase expression in clinical samples, we assessed BTL expression in different grades of clear cell kidney cell carcinoma (ccRCC).Results: Both new monoclonal antibodies bind to a protein in UOK171 cells but not in the negative control. Binding of mAb is specifc. mAb produced by cell line 2A9/2E9 (peptide 298–310; intracellular domain) is more suitable for immunohistochemical analyses as it gives stronger intensity of binding than mAb produced by cell line 11C9/2G9 (peptide 235–246; extracellular domain). Antibody 2A9/2E9 stains bilitranslocase in proximal renal tubules of normal kidneys but not in the surrounding stroma. Staining decreases in grade I compared to normal kidney, gradually increases in grades II and III, and decreases again in grade IV of ccRCC tissue.Conclusions: Our results show that these antibodies can be used in different immunoassays. Furthermore, specificity and afnity of our mAbs allowed us to use them in the analysis of progressive grades of clear cell renal cell carcinoma in a limited number of patients. Tus, mAbs developed here can be used as a diagnostic tool that could help distinguish between early and late grades of clear cell renal cell carcinoma.

Keywords

• clear cell renal cell carcinoma
• bilitranslocase
• monoclonal antibodies
• UOk171