Antibodies to the c3d fragment of complement component 3
一种抗体、补体的技术,应用在抗体、补体蛋白、抗体模拟物/支架等方向
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[0350] Anti-C3d antibody:
[0351] Recombinant human C3d was produced in E. coli using the pGEX expression system as described in Hannan, J.P., et al. (2005) J. Mol. Biol. 346, 845-858. Briefly, ampicillin-resistant clones were used to initiate overnight cultures, and cultures were expanded to 1 L and grown at 37°C until reaching A 600 =0.3. Cultures were induced with 0.3 mM isopropyl-β-D-thiogalactoside overnight at 30°C and then harvested by centrifugation. The harvested bacterial pellet was resuspended in glutathione-s-transferase column buffer (50 mM Tris-HCl, pH 8.0, 250 mM NaCl, 1 mM EDTA) and lysed by sonication. Lysates were clarified by centrifugation and applied to GStrap columns (GE Biosciences). C3d was cleaved from the column by digestion with 50 units of thrombin overnight at 4°C, followed by purification by size exclusion chromatography. The purity of C3d was monitored by SDS-PAGE.
[0352] C3 was immunized with 60 μg of recombinant C3d emulsified in Freund...
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