A thymosin beta 4 ELISA using an antibody against the N terminal fragment thymosin beta 4 [1-14].
A thymosin beta 4 ELISA was developed in which thymosin beta 4, absorbed on microwells, competed with thymosin beta 4 in solution for the binding sites of an anti-thymosin beta 4 antibody. The antibody molecules finally immobilized on the microwells were detected using a goat anti-rabbit immunoglobulin/horseradish peroxidase conjugate in combination with the substrate 2,2'-azino-bis-(3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt, and measuring the relevant optical density values. Anti-thymosin beta 4 antibodies were raised in rabbits against intact thymosin beta 4 as well as against selected fragments of the peptide, i.e., the N terminal fragments thymosin beta 4[1-14] and thymosin beta 4[1-11]. The antibody against thymosin beta 4[1-14] was used in the thymosin beta 4 ELISA, because it showed minimal cross-reactivity (0.1%) with the highly homologous peptide thymosin beta 9 as well as exhibiting the highest titre. The ELISA procedure developed, apart from showing a minimal cross-reaction with thymosin beta 9, was fast, easy to perform and exhibited good assay characteristics.
Livaniou
E
E
Abteilung für Physikalische Biochemie, Universität Tübingen, Germany.
Mihelic
M
M
Evangelatos
G P
GP
Haritos
A
A
Voelter
W
W
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D001667:Binding, Competitive; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D013947:Thymosin / Q000031:analogs & derivatives* / Q000032:analysis / Q000276:immunology; D000099201:Thymosin beta(4)
D000911:Antibodies, Monoclonal; D013947:Thymosin; D000099201:Thymosin beta(4); C042362:thymosin beta(9)