EPITOPE MAPPING AND BINDING ANALYSIS OF INSULIN-SPECIFIC MONOCLONAL-ANTIBODIES USING A BIOSENSOR APPROACH
EPITOPE MAPPING AND BINDING ANALYSIS OF INSULIN-SPECIFIC MONOCLONAL-ANTIBODIES USING A BIOSENSOR APPROACH
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DOI：
10.1016/0022-1759(95)00020-b
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发表时间：
1995-06-14
影响因子：
2.2
通讯作者：
BOUANANI, M
中科院分区：
文献类型：
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作者：
ALLAUZEN, S;MANI, JC;BOUANANI, M
用表面等离子体共振法研究了人胰岛素分子与一组抗胰岛素的单抗之间的分子相互作用。利用实时分析(Biacore技术)，绘制了这些单抗识别的人胰岛素分子上的抗原位点。根据与葡聚糖基质偶联的胰岛素分子被三个mAb群识别，可在其上定义三个重叠区域：组I(mAb19，10)、组II(mAb13，5，25，15)和组III(MAbP10)。该分子在溶液中的抗原决定簇使用两个位点的分析，其中胰岛素被呈递给每个单抗，结合到与葡聚糖偶联的兔抗鼠Ig G Fc Gamma 1。定义了四个抗原区：区I和区III由相同的抗体定义，而区II由单抗13、5(区IIa)和单抗25、15(区IIb)不同地识别。用Biaccore测定的单抗的K-a值在3×10(7)M(-1)到6.6×10(9)M(-1)的范围内，无论是胰岛素偶联到葡聚糖基质还是在溶液中。此外，选择识别胰岛素分子上两个不同区域的mAb19和P10，建立了一种测定血清中胰岛素的高灵敏免疫分析方法。
Molecular interactions between the human insulin molecule and a panel of seven monoclonal antibodies (mAbs) against insulin were investigated by surface plasmon resonance. Using real-time analysis (BIAcore technology), the antigenic sites recognized on the human insulin molecule by these mAbs were mapped. Three overlapping regions could be defined on the insulin molecule coupled to the dextran matrix on the basis of its recognition by three mAbs groups: group I (mAbs 19, 10), group II(mAbs 13, 5, 25, 15) and group III (mAb P10). The antigenic determinants of the molecule in solution were mapped using a two-site assay in which insulin was presented to each mAb, bound to a rabbit anti-mouse IgG Fc gamma 1 coupled to the dextran. Four antigenic regions were defined: regions I and III were defined by the same antibodies as above whereas region II was recognized differently by mAbs 13, 5 (region IIa) and mAbs 25,15 (region IIb). The K-a value of the mAbs determined using the BIAcore were in the same range (from 3 x 10(7) M(-1) to 6.6 x 10(9) M(-1)) whether insulin was coupled to the dextran matrix or in solution. Furthermore, mAbs 19 and P10, which recognize two distinct regions on the insulin molecule, were selected to develop a highly sensitive immunoassay for measurement of insulin in serum.