Evaluation of progesterone-ovalbumin conjugates with different length linkers in enzyme-linked immunosorbant assay and surface plasmon resonance-based immunoassay

• Published in: Steroids 2002-06, Vol.67 (7), p.565-572
• Main Authors: Wu, Yinqiu, Mitchell, John, Cook, Christian, Main, Lyndsay
• Format: Article
• Language: English
• Subjects: Anti-progesterone mAb; BIAcore; Biological and medical sciences; Carboxylic Acids - chemical synthesis; Carboxylic Acids - chemistry; ELISA; Enzyme-Linked Immunosorbent Assay - standards; Fundamental and applied biological sciences. Psychology; Molecular Structure; Ovalbumin - chemistry; Ovalbumin - immunology; Particle Size; Progesterone - chemistry; Progesterone - immunology; Progesterone-OVA conjugates; Reference Standards; Steroid; Steroid hormones. Cholecalciferol derivatives; Surface Plasmon Resonance; Vertebrates: endocrinology
• ISSN: 0039-128X; 1878-5867
• DOI: 10.1016/S0039-128X(02)00010-7

A series of progesterone-4-ovalbumin (OVA) conjugates with different length linkers (4-, 11-, and 18-atoms long) were synthesized by successive aminocaproic acid homologation of 3-(pregn-4-ene-3,20-dione-4-yl)thiopropanoic acid ( 1) before conjugation to ovalbumin. The performance studies of these progesterone-4-ovalbumin conjugates showed that the effects of the length of linker on the antibody binding are dependent upon different immunoassay formats. In a rapid flow biosensor surface, on a BIAcore Surface Plasmon Resonance (SPR) instrument, antibody-binding capacities and response rate were dramatically increased for progesterone-4-ovalbumin conjugates when the length of the linker was incremented from 4 atoms to 11 or 18 atoms. Thus, highly sensitive SPR-based immunoassays for progesterone over a range of 0.1–50 ng ml −1 were developed using biosensor surfaces immobilized with progesterone-ovalbumin conjugates having extended linkers. The SPR-based assays were fully competitive with conventional enzyme-linked immunosorbant assay (ELISA) but much more rapid and simple. However, there were little changes in antibody-binding performance using a conventional ELISA for the same conjugates. The progesterone-4-ovalbumin conjugate ( 1-OVA) had better antibody binding than its progesterone-7α-ovalbumin analog ( 2-OVA) in the SPR-based assay, but with a conventional ELISA there was no significant difference between these two isomeric conjugates.