Effect of an Exposure to Chloramine-T on the Immunoreactivity of Glucagon¹

Abstract

Various preparations of glucagon treated with chloramine-T under different conditions have been studied with respect to their immunoreactivity toward two different glucagon antisera; one specific for pancreatic glucagon and the other capable of reacting with enteroglucagon as well. The glucagon preparations exposed to chloramine-T for different periods reacted almost identically with the nonspecific antibody whether they were used as tracer or standard. On the contrary, treatment with chloramine-T under severe conditions led to reduced immunoreactivity toward the specific antibody. Inclusion of dimethyl sulfoxide (DMSO) in the chloramine-T reaction resulted in preservation of the immunoreactivity of the treated preparations. The cyanogen bromide cleaved-glucagon, (1-26) homoserine lactone, showed little cross-reactivity with the specific antibody whereas it reacted to a similar extent with the nonspecific antibody as natural glucagon did. Amino acid analysis of the hormone exposed to chloramine-T demonstrated that the methionine residue at position 27 in the glucagon molecule had been oxidized to methionine sulfoxide. In addition, tryptophan had also been affected. DMSO protected methionine and tryptophan from the oxidative action of chloramine-T. We postulate from these results that the change in the immunoreactivity toward the specific antibody of glucagon exposed to chloramine-T is mainly due to oxidation of the methionine residue at position 27 in the molecule. The usefulness of DMSO in the iodination process is also discussed.

Lack of gastrointestinal glucagon response to hypoglycaemia in depancreatized dogs

“…GI was measured with a specific antiserum for "pancreatic" glucagon reacts with gut extract, to assay total glucagon-like immunoreactivity (total GLI), and AGS 18 which is a so-called specific antiserum for "pancreatic" glucagon (GI). The characteristics of these antisera have been discussed elsewhere ]1, 6,8]. In addition they may be characterised, as follows: AGS 10 is apparently not Cterminal specific but it is yet uncertain exactly at which position it binds.…”

“…In addition they may be characterised, as follows: AGS 10 is apparently not Cterminal specific but it is yet uncertain exactly at which position it binds. AGS 18 is specific for the C-terminus of the glucagon molecule or the 18-29 region including the methionine residue at position 27 [8]. Gut glucagon-like immunoreactivity (gut GLI) was obtained by subtracting GI from total GLI.…”

Glucagon-like polypeptide and insulin contents in the brain from acute hepatic failure dogs

“…All the brain fragments were immediately extracted with 0.2N acetic acid according to the method of Tanaka et al [14]. The extracts were reconstructed with water and then measured by RIA using C-terminal specific glucagon antiserum, OAL-123, for GI contents, and N-terminal specific glucagon antiserum, OAL-196, for total GLI levels [11]. GLI contents were calculated by subtracting GI contents from total GLI contents [11].…”

“…The extracts were reconstructed with water and then measured by RIA using C-terminal specific glucagon antiserum, OAL-123, for GI contents, and N-terminal specific glucagon antiserum, OAL-196, for total GLI levels [11]. GLI contents were calculated by subtracting GI contents from total GLI contents [11]. GI and GLI contents in the brain were expressed as pg of g brain wet weight.…”

Comparison of the Changes in Plasma Human Growth Hormone (HGH) and Immuno-Reactive Glucagon (IRG) after Intravenous and Subcutaneous Injection of Glucagon

“…HGH was assayed by the doubleantibody radioimmunoassay (Schalch and Parker, 1964) using Wilhelmi HGH as the standard, the lower limit of sensitivity was 1ng/ml. Glucagon (IRG) was measured by the radioimmunoassay method described previously (Shima et al, 1975) using nonspecific antiglucagon serum. Plasma FFA was measured spectrometrically (Duncombe, 1964) and plasma lactate was determined by the method reported by Scholz et al (1959).…”