Analysis of the interaction between monoclonal antibodies and human hemoglobin (native and cross-linked) using a surface plasmon resonance (SPR) biosensor.
To develop a stable immuno-assay system for quantification of human hemoglobin (Hb), the interaction between various antibodies and Hb was studied using a surface plasmon resonance (SPR) biosensor in the BIAcore equipment (Amersham Pharmacia Biotech) with an immobilized anti-Hb antibody sensor chip. When polyclonal antibodies were used, the immuno-reactivity of purified and commercially available Hb decreased drastically with incubation times up to 14 h. This instability of immuno-reactivity of Hb is attributable to the conformational changes in Hb induced by oxidation. On the other hand, of the sixteen monoclonal antibodies tested, four antibodies (MSU-102, -103, -106 and -115) were found to maintain their immuno-reactivities at least up to 24 h. During long-term storage, however, the immuno-reactivity of Hb with these monoclonal antibodies decreased significantly. The chemical betabeta-cross-linking of Hb was effectively able to stabilize the structure of Hb and immuno-reactivity with monoclonal antibodies such as MSU-103 for periods at least up to 70 days. Therefore, the combination of specific monoclonal antibodies such as MSU-103 and a betabeta-cross-linked Hb standard could be used for the quantification of Hb.
Sonezaki
S
S
Kokura R&D Division, R&D Center, TOTO Ltd., 1-1 Nakashima, 2 Kokurakitaku, Kitakyushu, Japan. shuji.sonezaki@toto.co.jp
Yagi
S
S
Ogawa
E
E
Kondo
A
A
D000911:Antibodies, Monoclonal / Q000276:immunology*; D015374:Biosensing Techniques; D002138:Calibration; D003432:Cross-Linking Reagents; D006454:Hemoglobins / Q000276:immunology*; D006801:Humans; D020349:Surface Plasmon Resonance / Q000379:methods; D013997:Time Factors
D000911:Antibodies, Monoclonal; D003432:Cross-Linking Reagents; D006454:Hemoglobins