Editing SPR

Surface plasmon resonance (SPR)<br/>For the immobilization onto CM5 chip, all antigens were<br/>activated via thiol labeling kit (GE Healthcare, Buckinghamshire,<br/>UK). A 0.5 mg of antigens in 0.5 mL were prepared in<br/>morpholinoethanesulfonic acid (MES) buffer. Then, 0.25 mL of<br/>2-(2-pyridinyldithio)ethaneamine (PDEA) (15 mg/mL) in 0.1 M<br/>MES buffer was added. After adding 25 mL of 0.4 M 1-ethyl-3-(3-<br/>dimethylaminopropyl)-carbodiimide (EDC), the mixtures were<br/>incubated at 25uC for 10 min. The buffers were exchanged to PBS<br/>via dialysis after the incubation. A Biacore 3000 (GE Healthcare)<br/>and CM5 chip (GE Healthcare) was used in all SPR analysis. SPR<br/>experiments were carried out at a flow rate of 10 mL/min using<br/>PBS as running buffer. For the activation of the surface of CM5<br/>chip, 0.2 M EDC and 0.5 M N-hydroxysuccinimide (NHS)<br/>solution was injected for 2 min. And 40 mM cystamine solution<br/>was injected for 3 min, and then, 0.1 M dithioerythritol (DTE) in<br/>0.1 sodium borate solution was injected for 3 min. After the<br/>activation of the chip surfaces, 100 mg/mL of activated antigens<br/>were injected for 7 min. For deactivation of excess reactive groups<br/>on the chip surface, 20 mM PDEA and 1 M NaCl solution in<br/>0.1 M sodium acetate was injected for 4 min. Finally, various<br/>concentrations of isolated scFvs were injected for 5 min and the<br/>binding signal was detected. The KD values of scFvs were<br/>evaluated by measuring Kon and Koff in BIA evaluation software
[[Category:Antibodies]]