Transition metal carbonyl labeling of proteins. A novel approach to a solid-phase two-site immunoassay using Fourier transform infrared spectroscopy.
Labeling of bovine serum albumin (BSA) and anti-human thyroid stimulating hormone (hTSH) monoclonal antibodies (mAbs) was performed using (N-succinimidyl 4-pentynoate)hexacarbonyldicobalt (NSCo2(CO)6). Conditions of coupling were different depending on the protein to be labeled, denaturation of the mAbs occuring with high percentages of organic solvent in the reaction mixture. The influence of reaction time and initial concentration of NSCo2(CO)6 was examined. They were both shown to affect the final coupling rate of the metal carbonyl probe. Preservation of the immunoreactivity toward 125I-hTSH was observed for five conjugates having different NSCo2(CO)6: mAb molar ratios when compared to unmodified and peroxidase-labeled mAbs. Finally, a preliminary study of the quantitative detection of the metal carbonyl mAbs on microtiter wells was achieved using Fourier transform infrared spectroscopy.
Varenne
A
A
Ecole Nationale Supérieure de Chimie de Paris, France.
Salmain
M
M
Brisson
C
C
Jaouen
G
G
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology; D002417:Cattle; D003035:Cobalt / Q000737:chemistry*; D005583:Fourier Analysis; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D007202:Indicators and Reagents; D015335:Molecular Probes; D009942:Organometallic Compounds / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000737:chemistry; D013055:Spectrophotometry, Infrared; D013388:Succinimides / Q000737:chemistry*; D013972:Thyrotropin / Q000276:immunology
D000911:Antibodies, Monoclonal; D007202:Indicators and Reagents; D015335:Molecular Probes; D009942:Organometallic Compounds; D013388:Succinimides; C069220:(N-succinimidyl 4-pentynoate)hexacarbonyldicobalt; D012710:Serum Albumin, Bovine; D003035:Cobalt; D013972:Thyrotropin