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Immunofluorescence to Monitor the Cellular Uptake of Human Lactoferrin and its Associated Antiviral Activity Against the Hepatitis C Virus
Published on: October 1, 2015
[Isolation and purification of recombinant human lactoferrin (rhLF) from transgenic rice and its antibacterial
Jinghuan Wu1, Lichen Yang, Gaige Liu
1Institute of Nutrition and Food Safety, Chinese Centre for Disease Control and Prevention, Beijing 100050, China. jhwu2012@163.com
Objective:
To study the isolation and purification of recombinant human lactoferrin (rhLF) from transgenic rice, and to check its antibacterial activities.
Methods:
After isolated rhLF from transgenic rice via saturated ammonium sulfate precipitation, then purified it through CM Sepharose FF-exchange chromatography and molecular sieve chromatography Sephadex G25. The inhibition effects under different concentrations of rhLF (0.25, 0.5, 1.0, 2.0, 4.0, and 5.0 mg/ml) against Salmonella typhimurium, Staphyloccocus aureus, Bacillus cereus, Listeria monocytogenes were observed, using broth microdilution method.
Results:
The rhLF was obtained at a higher purity (about 90%) through successful isolation and purification. After Coomassie blue staining, Westernblot and mass spectrometer analysis, it was identified as the purpose protein with the molecular weight of approximately 79 kDa. The antibacterial experiments showed that 5 mg/ml and 4 mg/ml rhLF could inhibite Salmonella typhimurium and Staphylococcus aureus persistently, 2 mg/ml and 1 mg/ml rhLF showed a significant inhibitory effects in the later period; while 0.5 mg/ml or lower concentration, showed no inhibitory effects. As to Bacillus cereus, only 5 mg/ml and 4 mg/ml rhLF exhibited certain inhibitory effects within 18 hours. Listeria monocytogenes was inhibited within 18 hours just at 5 mg/ml rhLF.
Conclusion:
The rhLF could be successfully separated and purified from transgenic rice, and the purified protein still has significant antibacterial activities.
