We produced three monoclonal antibodies against different epitopes of the precursor form of atrial natriuretic polypeptide (ANP), γ -human ANP (γ-hANP) [human atrial natriuretic factor (ANF)-(1-126)]. Monoclonal antibodies were prepared by fusion of mouse myeloma cells X63-Ag8.653 with spleen cells from BALB/c mice immunized with synthetic α-hANP or γ -hANP-(1-25) [human ANF-(1-25)] conjugated to thyroglob- ulin. Of three monoclonal antibodies obtained, two were directed against the α -ANP sequence: one antibody (KY-ANP-I) recognizes the N-terminal half of the ring structure, while the other (KY-ANP-II) recognizes the N-terminal sequence of α -hANP. The third antibody (KY-ANP-III) was against γ -hANP-(1-25). With the aid of rabbit polyclonal anti-α-ANP-(17-28), we developed sandwich enzyme immunoassays. An enzyme immunoassay for a-hANP using KY-ANP-I and polyclonal Fab’ was very sensitive (0.01 fmol/tube) and we could detect 0.6 pg/ml of plasma α -hANP without extraction. Thus, monoclonal antibodies against γ -ANP are useful tools for investigating the significance of ANP and related peptides derived from γ-ANP.
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