Evaluation of progesterone-ovalbumin conjugates with different length linkers in enzyme-linked immunosorbant assay and surface plasmon resonance-based immunoassay.
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-7alpha-ovalbumin analog (2-OVA) in the SPR-based assay, but with a conventional ELISA there was no significant difference between these two isomeric conjugates.
Wu
Yinqiu
Y
BioEngineering Technologies, HortResearch, East Street, Private Bag 3123, Hamilton, New Zealand. ywu@hortresearch.co.nz
Mitchell
John
J
Cook
Christian
C
Main
Lyndsay
L
D002264:Carboxylic Acids / Q000138:chemical synthesis / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay / Q000592:standards; D015394:Molecular Structure; D010047:Ovalbumin / Q000737:chemistry* / Q000276:immunology*; D010316:Particle Size; D011374:Progesterone / Q000737:chemistry* / Q000276:immunology*; D012015:Reference Standards; D020349:Surface Plasmon Resonance
D002264:Carboxylic Acids; D011374:Progesterone; D010047:Ovalbumin