抗原，抗体（鼠，兔）中二者相互作用检测方案(大分子作用仪)

HORIBAJOBINYVONSPRi & Life Sciences SPRi & Life SciencesApplication Note SPRi-0707.2-v1- Page 2 Antibody-Antigen Specific Interaction GenOptics provides the pharmaceutical industry， biotech companies and academic research laboratories withhigh performance instrumentation based on SPRi technology (Surface Plasmon Resonance imaging). Todemonstrate the power of this technology， we carried out an experiment based on an antibody-antigeninteraction. Original surface chemistry combined with an electrochemical process allows the rapid coupling ofbiomolecules to the gold layer (polypyrrole co-polymerisation) on the top of a glass prism. Experiment Principle Two types of antibodies are first coupled to the pyrrolegroup and then grafted by electro-polymerisation onto theglass prism. -Rabbit IgG (blue)， -Anti-hCG antibodies from mice (red). Next， anti-rabbit lgG are added. Fig. 1：Layout of a 25 spots matrix composed of rabbit lgG and anti-hCC IgGspots Interactions between rabbit IgGand anti-rabbit lgG from goat In the first experiment， anti-lgG from rabbit are injected.Figure 2 represents the kinetics of interaction between theantibodies immobilized on the gold surface and the anti-rabbit lgG injected in the detection cell. There is a clear interaction between the rabbit lgG (redcurves) and injected antibodies. There is no reaction withother spots. The interaction rabbit lgG and anti-rabbit lgGfrom rabbit is specific and can be monitored on a real timebasis. Fig. 2： Reflectivity variations as a function of timefollowing injection of anti-lgG from rabbit A：PBS/Tween buffer B： Injection of the anti-lgG C： Rinsing with PBS/Tween buffer Example of interactions： hCG-mouse anti-hgG，hCG -rabbit anti-hCG，rabbit anti-hCG-anti-rabbit lgG In a second experiment， a mouse anti-hCG antibody isfirstly immobilised on the gold surface of the glass prism.The surface is then successively exposed to varioussolutions containing hCG， anti-hCG antibodies (differentfrom those already immobilised)， and a rabbit anti-lgGantibody. Fig. 3： Interactions between a mouse anti-hCC antibody and hCC (D)，then between hCC and a rabbit anti-UCC (F)， and between a rabbitantibody and an anti-rabbit lgG antibody(H). Thegraph below showsthesedifferent interactionsmonitored in real time： Fig. 4： Kinetic curves on mouse anti-hCG spots， rabbit lgG and bare surface， following subsequent injections of hCG， rabbit anti-hCG and rabbitanti-/gG. E， C， l： Injection of PBS 10 mM， Tween 0.05 DInjection of 250 nM hCG Injection of anti-hCG (monoclonal antibody 210 nM)， Injection of goat anti-rabbit antibody (polyclonal 220 nM) Conclusion This technique is thtehrerefefororee proven to be efficientt aatperforming accurate high throughput antibody screening. France： +33 (0)16454 13 00USA： +1 732 49486 60 Japan： +81 (0)33861 8231Germany： +49 (0)89 46 23 17-0UK：+44(0)2082048142 ltaly： +39 0 2 5760 3050China： +86(0)108567 9966Other Countries： +33 (0)1 64 54 13 00 HORIBAExplore the future Find us at www.jobinyvon.com/SPRi or telephone：HORIBAExplore the future GenOptics SPRi technology allows a real time study of biomolecular interactions. No additional labelling reagent is required, and a large number of spots can be analysed at the same time and in the same condition.This technique is therefore proven to be efficient at performing accurate high throughput antibody screening.