Studies on the Interaction between Biomolecules by Surface Plasmon Resonance Technology and Fluorescence Spectrometry
|Keywords||Surface plasma resonance Fluorescence spectrometry Antibody Drug protein|
Chapter 1:The principle, characteristics, research contents and applications of surface plasma resonance(SPR) technology had been reviewed. The BIAcore X SPR biosensor including its composition and characteristics were also introduced in detail. The fluorescence spectrometry and its research contents on the interaction between drugs and biological macromolecules were outlined as well. On the basis of these views, our own research contents were put forward.Chapter 2:The interaction of human immunoglobulin G (human IgG) and rabbit anti-human IgG antibody was investigated using SPR technology. The factors, including pH value, ionic strength, protein concentration, influencing electrostatic adsorption of ligand onto the carboxylated dextran-coated sensor chip surface were considered. The human IgG was immoblized on the sensor chip and the immune reaction was monitored in real time. The regeneration condition was also investigated. According to the sensorgram, the binding dynamic constant Ka=9.37×104L·mol-1·s-1 and affinity constant KD=4.09×10-8 mol·L-1 had been obtained by the evaluation software.Chapter 3:Competitive inhibition analysis was applied to study the interaction between chloramphenicol and its antibody by SPR technology. The chloramphenicol-BSA conjugate was immoblized onto the sensor chip, then the mixture of antibody and chloramphenicol were flowed past the surface of chip as analytes. The procedure of interaction was monitored in real time. In the detection range of 0-20ng/mL, the standard curve was drawed.Chapter 4:The interaction of VB12 and bovine serum albumin(BSA) were studied by fluorescence spectrometry. Fluorescence static quenching mechanism of BSA by VB12 was discussed. The association constant was KA =1.00×105L/mol. The number of binding site was 1.07. The binding distance was a distance of 6.29nm away from 212 tryptophan residue in BSA according to Forster’s non-radiative energy transfer mechanism.We also studied the effect of VB12 to tyrosine, tryptophan residues by synchronous fluorescence.Chapter 5:The interaction between prulifloxacin and BSA were studied by fluorescence spectrometry. Fluorescence quenching mechanism of bovine serum albumin by prulifloxacin was discussed. The assoation constant was KA=3.53×106L mol-1. The number of binding site was 1.32. The binding distance was a distance of 3.41nm away from tryptophan residue in BSA according to Forster’s non-radiative energy transfer mechanism.