Allauzen S, Mani J C, Granier C, Pau B, Bouanani M
CNRS UMR 9921, UFR Pharmacie, Montpellier, France.
J Immunol Methods. 1995 Jun 14;183(1):27-32. doi: 10.1016/0022-1759(95)00020-b.
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 Ka 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.
利用表面等离子体共振技术研究了人胰岛素分子与一组七种抗胰岛素单克隆抗体（mAb）之间的分子相互作用。通过实时分析（BIAcore技术），绘制了这些单克隆抗体在人胰岛素分子上识别的抗原位点。基于三组单克隆抗体对结合到葡聚糖基质上的胰岛素分子的识别，可在胰岛素分子上定义三个重叠区域：第一组（单克隆抗体19、10）、第二组（单克隆抗体13、5、25、15）和第三组（单克隆抗体P10）。使用双位点分析法绘制溶液中该分子的抗原决定簇，其中将胰岛素呈递给每种单克隆抗体，该单克隆抗体与结合到葡聚糖上的兔抗小鼠IgG Fcγ1结合。定义了四个抗原区域：区域I和III由上述相同抗体定义，而区域II被单克隆抗体13、5（区域IIa）和单克隆抗体25、15（区域IIb）以不同方式识别。无论胰岛素是偶联到葡聚糖基质上还是处于溶液中，使用BIAcore测定的单克隆抗体的Ka值都在相同范围内（从3×10⁷ M⁻¹到6.6×10⁹ M⁻¹）。此外，选择识别胰岛素分子上两个不同区域的单克隆抗体19和P10，开发一种用于测定血清中胰岛素的高灵敏度免疫测定法。
