Evaluation on effect of ultrafiltration technology on endotoxin removal from recombinant MUC1-MBP fusion protein
10.13481/j.1671-587x.20140315
- VernacularTitle:超滤技术去除重组MUC1-MBP融合蛋白中内毒素的效果评价
- Author:
Juan WANG
;
Fei XIE
;
Tanxiu CHEN
;
Xiaxia SUN
;
Qiongshu LI
;
Guixiang TAI
- Publication Type:Journal Article
- Keywords:
MUC1-MBP fusion protein;
ion exchange chromatography;
thin layer chromatography;
ultrafiltration technology;
endotoxin;
chromogenic end-point tachypleus amebocyte lysate
- From:
Journal of Jilin University(Medicine Edition)
2014;(3):539-542
- CountryChina
- Language:Chinese
-
Abstract:
Objective To evaluate the effectiveness of ultrafiltration technology in endotoxin removal from purified recombinant MUC1-MBP fusion protein (MUC1-MBP)and to demonstrate the effect of ultrafiltration on endotoxin removal.Methods CM Sepharose FF weak cation exchange (CM)(CM group), CM combined with Phenyl Sepharose 6 FF exchange (C6)(CM+C6 group),CM combined with ultrafiltration (CM+ultrafiltration group), and CM combined with C6 and ultrafiltration (CM+C6+ultrafiltration group)were used to purify the MUC1-MBP from E.coli. and remove endotoxin;the expression level of endotoxin was detected by Chromogenic End-point Tachypleus Amebocyte Lysate.Results There was a single band at the expected molecular weight of 62 000 by SDS-PAGE analysis.and the purity>96% by Quantity One analysis.The endotoxin levels in CM group and CM +C6 group were quite high and there was no significant difference between two groups (P>0.05 );the endotoxin level in CM+ultrafiltration group was significantly lower than that in CM group, and there was significant difference (P<0.01);the endotoxin level in CM+C6+ultrafiltration group was significantly decreased compared with CM+C6 group (P<0.01);there was no significant differences of endotoxin level between CM+ultrafiltration group and CM +C6+ultrafiltration group (P>0.05 ). Conclusion The effects of CM or CM combined with C6 on endotoxin removal are quite poor, especially C6;CM combined with ultrafiltration are quite effective on endotoxin removal,and ultrafiltration plays an important role in endotoxin removal.
