Production and evaluation of secretin antibodies

Abstract

Antibodies were readily produced in three rabbits to unconjugated pure natural porcine secretin, and in two rabbits to synthetic porcine secretin conjugated to BSA. The final dilutions of the antisera to bind 50% of 1 fmol 125-i-labeled secretin prepared by the Chloramine-T method and purified on a Sephadex G-15 and a SP Sephadex C-25 column varied between 1:14,500 and 1:245,000. The effective equilibrium constants (Keff) according to Scatchard varied between 0.8 x 10(11) and 3.4 x 10(11), the average equilibrium constants (Ko) according to Sips varied between 0.8 x 10(11) and 3.6 x 10(11), and the indices of heterogeneity (alpha) according to Sips were 1.00 or close to 1.00. None of the antisera showed any cross reactivity to gastrin, glucagon or insulin. There was no differences in any of the preceding parameters between the antisera produced to the two immunogen preparations. It is suggested that the immunogenicity of secretin may be related to its basic charge, and to the possibility that secretin may circulate bound to certain plasmafactors or in a polymerized form. The two antisera with the highest equilibrium constants, which also allowed the highest working dilution to be applied, allowed measurements of fasting plasma secretin levels in the low pmol/1 range in all acidified plasmas examined.