Use of antibodies against synthetic peptides for analysis of molecular multiplicity: human deoxyribonuclease I polymorphism.
Eight different antibodies were raised in rabbits and chickens by injecting them with two synthetic N- and C-peptides, which corresponded to the N- and C-terminal 15 residues of human deoxyribonuclease I (DNase I). These two peptides were used as immunogens, both free and conjugated with keyhole limpet hemocyanin (KLH). A rabbit antiserum against the C-peptide conjugated with KLH (rabbit anti-C/KLH) and two chicken antisera against the C-peptide itself (chicken anti-C) and conjugated with KLH (chicken anti-C/KLH) reacted well with purified DNase I, blotted onto a transfer membrane after isoelectric focusing in a polyacrylamide gel. A clear isoenzyme pattern identical to that detected with a rabbit antiserum against the parent enzyme (rabbit anti-DNase I) was observed. Surprisingly, the chicken anti-C antiserum was found to be a powerful and highly specific tool for the determination of urinary DNase I phenotypes (Kishi et al., Hum. Genet. 1989, 81, 295-297) and was not inferior in any respect to the rabbit anti-DNase I. These findings show that the selection of immune animal is one of the most important factors for producing an effective anti-peptide antibody. Unfortunately, the anti-peptide antisera obtained in this study neither blocked DNase I enzyme activity nor did it bind to the enzyme.
Yasuda
T
T
Department of Legal Medicine, Fukui Medical School, Japan.
Nadano
D
D
Takeshita
H
H
Kishi
K
K
D000818:Animals; D000918:Antibody Specificity; D002645:Chickens; D003429:Cross Reactions; D003850:Deoxyribonuclease I / Q000032:analysis / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007106:Immune Sera; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000032:analysis / Q000235:genetics*; D010455:Peptides / Q000032:analysis*; D011110:Polymorphism, Genetic; D011817:Rabbits; D013194:Staining and Labeling
D007106:Immune Sera; D007527:Isoenzymes; D010455:Peptides; D003850:Deoxyribonuclease I