Agarose hydrophobic chromatoghaphy medium and its application in purifying yeast expression HBsAg
A technology of hydrophobic chromatography and agarose, applied in the biological field, can solve problems such as uncontrollable ligand density, weakened adsorption capacity of biological macromolecular media, and failure of chromatography media to meet application requirements, etc., to achieve easy maintenance of biological activity and excellent biological Compatibility, the effect of simple and easy purification method
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Embodiment 1
[0040] Take 100.0 g of washed and drained agarose 6FF gel (manufactured by Hangzhou Zhengguang Resin Co., Ltd.), and pour it into a 1-liter three-necked flask. To this was added 150 mL of acetone, 75 mL of epichlorohydrin and 75 mL of 0.5M NaOH solution. The reaction was stirred at 60°C for 4 hours. After the reaction, the gel was washed with water and vacuum-dried, and the washing process was repeated three times. Take 100.0 g of agarose epoxy-activated gel that has been washed and drained, and pour it into a 1-liter three-necked flask. 100 μL of butanethiol, 10 mL of ethanol, and 90 ml of 0.1 M NaOH solution were added thereto. Stir at 60°C for 6 hours. After the reaction, the gel was washed with water and dried, and the washing process was repeated three times to obtain the agarose hydrophobic chromatography medium of the present invention, with a ligand density of 100 μmol / g dry gel, stored in 20% ethanol solution.
Embodiment 2
[0042] Take 100.0 g of washed and drained agarose 6FF gel and pour it into a 1-liter three-necked flask. To this was added 150 mL of epichlorohydrin and 75 mL of 4M NaOH solution. The reaction was stirred at 25°C for 10 hours. After the reaction, the gel was washed with water and dried, and the washing process was repeated three times. Take 100.0 g of agarose epoxy-activated gel that has been washed and drained, and pour it into a 1-liter three-necked flask. 2000 μL of butanethiol, 10 mL of ethanol and 90 ml of 4M NaOH solution were added thereto. Stir at 25°C for 10 hours. After the reaction, the gel was washed with water and dried, and the washing process was repeated three times to obtain the agarose hydrophobic chromatography medium of the present invention with a ligand density of 120 μmol / g dry gel, which was stored in 20% ethanol solution.
Embodiment 3
[0044] Take 100.0 g of washed and drained agarose 6FF gel and pour it into a 1-liter three-necked flask. Thereto were added 150 mL of acetone, 75 mL of 1,4-butanediol diglycidyl ether and 75 mL of 0.5M NaOH solution. The reaction was stirred at 60°C for 4 hours. After the reaction, the gel was washed and dried, and the washing process was repeated three times. Take 100.0 g of agarose epoxy-activated gel that has been washed and drained, and pour it into a 1-liter three-necked flask. 100 μL of butanethiol, 10 mL of ethanol and 90 ml of 0.1 M NaOH solution were added thereto. Stir at 60°C for 6 hours. After the reaction, the gel was washed and dried, and the washing process was repeated three times to obtain the agarose hydrophobic chromatography medium of the present invention with a ligand density of 61 μmol / g dry gel, which was stored in 20% ethanol solution.
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