ABSTRACT
Plant bioreactor is a new production platform for expression of recombinant protein, which is one of the cores of molecular farming. In this study, the anti DYKDDDDK (FLAG) antibody was recombinantly expressed in tobacco (Nicotiana benthamiana) and purified. FLAG antibody with high affinity was obtained after immunizing mice for several times and its sequence was determined. Based on this, virus vectors expressing heavy chain (HC) and light chain (LC) inoculated into Nicotiana benthamiana leaves by using Agrobacterium-mediated delivery. Accumulation of the HC and LC was analyzed by SDS/PAGE followed by Western blotting probed with specific antibodies from 2 to 9 days postinfiltration (dpi). Accumulation of the FLAG antibody displayed at 3 dpi, and reached a maximum at 5 dpi. It was estimated that 66 mg of antibody per kilogram of fresh leaves could be obtained. After separation and purification, the antibody was concentrated to 1 mg/mL. The 1:10 000 diluted antibody can probe with 1 ng/mL FLAG fused antigen well, indicating the high affinity of the FLAG antibody produced in plants. In conclusion, the plant bioreactor is able to produce high affinity FLAG antibodies, with the characteristics of simplicity, low cost and highly added value, which contains enormous potential for the rapid and abundant biosynthesis of antibodies.
Subject(s)
Animals , Mice , Antibodies , Nicotiana/genetics , Agrobacterium/genetics , Bioreactors , Blotting, WesternABSTRACT
Objective To investigate the effect of total glucosides of paeong (TGP) on the liver lipid infiltration and fibrosis in rats with non-alcoholic fatty liver disease (NAFLD) induced by fructose and high-fat diet. Methods Fructose-high-fatty induced NAFLD rat model was established. Metformin ( MET,200 mg · kg-1 ) and TGP (200,100 mg · kg-1 ) was intragastrically given to the rats in the treatment group,TGP high dose and low dose group,respectively. Normal control group and model control group was intragastrically treated with equivalent distilled water (10 mL·kg-1 ). At the fourth week after the treatment,all the rats were sacrificed and the indices such as serum fasting blood glucose(FBG),INS,insulin sensitivity index (ISI),triglycerides(TG),apelin-36,visfatin,alanine aminotransferase(ALT),aspartate aminotransferase(AST),free fatty acid (FFA),collagen Ⅲ(COLⅢ),collagen Ⅳ(COLⅣ) were determined. Hepatic content of TG was determined and the pathological changes in the liver tissues were observed under the microscope. Results As compared with the model control group,TGP effectively decreased FBG,INS,TG in serum and liver tissues,activity of ALT and AST in serum and content of FAA,Apelin, Visfatin,COLⅢ and COLⅣ,with significant differences (P<0. 05 or P<0. 01). TGP alleviated lipid infiltration and fibrosis in rat liver tissues. Conclusion TGP can inhibit effectively lipid infiltration and fibrosis of NAFLD rats,probably through improving glucolipid metabolism and antogonizing insulin resistance.
ABSTRACT
OBJECTIVE: To establish a bacterial endotoxin test method for fluorescein sodium injection.METHODS: Based on the bacterial endotoxin test method stated in Chinese Pharmacopoeia 2005 edition(second part),an interference test was conduced on different batches of fluorescein sodium samples using tachypleus amebocyte lysate from different companies.RESULTS: At a concentration of no more than 0.75 mg?mL-1,fluorescein sodium showed no interfere with the bacterial endotoxin test.CONCLUSION: It is feasible for bacterial endotoxin test of fluorescein sodium injection be conducted in a limit value of 0.15 EU?mL-1.