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Separation purification and identification methods of chicken apolipoprotein A II

A technology for separation and purification of apolipoprotein AII, which is applied in the field of separation, purification and identification of chicken apolipoprotein AII, can solve the problems of incomplete elucidation of physiological functions and obstruction of intracellular cholesterol transport, etc., and achieves strong practicability, simple method, The effect of easy operation

Inactive Publication Date: 2014-12-31
胡国良
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Problems solved by technology

[0003] The physiological function of ApoAⅡ has not been fully elucidated so far. In addition to being involved in maintaining the structure of HDL, it may also have the function of inhibiting the activity of lecithin cholesterol acyltransferase (LCAT), thereby hindering the export, esterification and transfer of cholesterol in cells.

Method used

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Embodiment Construction

[0019] This specific embodiment adopts the following technical solutions, and its separation and purification method is:

[0020] 1 Collection of chicken serum: blood is collected from the vein under the wing. After the blood is collected, remove the needle and gently inject it into the test tube. In order to prevent the rupture of blood cells, cleaned and sterilized syringes and test tubes are infiltrated with 0.85% normal saline in advance to prevent blood clots from adhering to the glass tube wall, resulting in poor serum precipitation. The test tube is placed in a 37°C incubator at a 45° angle for 1 hour, and then placed in a 4°C refrigerator for 3 to 4 hours. After the blood clot shrinks, the serum is drawn with a capillary dropper, and then centrifuged at 1000×g for 10 min. After clearing the serum, add NaN in the refrigerator at -20℃ 3 Save it for later use.

[0021] 2 Preparation of HDL crude product-phosphotungstic acid precipitation method: take 1.0 ml of serum and add 1...

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Abstract

The invention discloses separation purification and identification methods of a chicken apolipoprotein A II, and relates to the technical field of biology. The separation purification method comprises chicken serum collection, crude HDL preparation, HDL crude product degreasing, degreased HDL dissolving, dialysis and concentration of the degreased HDL solution, extraction of the apolipoprotein A II, chromatography with a DEAESepharoseCL-6B column, and chromatography with a SephadexG-150 column; according to the method, the SephadexG-150 chromatography column is prepared, then an apoA II-containing protein solution passing through the DEAESepharoseCL-6B is added on a gel column, when the protein solution is parallel and level to a colloid, elution is carried out by using an eluant, concentration is carried out with sucrose, then the product is preserved for standby analysis. The method is simple, is strong in practicability, helps people further understand physiological functions of the apoA II, and can be used in epidemiological and clinical studies.

Description

Technical field: [0001] The invention relates to the field of biotechnology, in particular to a method for separating, purifying and identifying chicken apolipoprotein AII. Background technique: [0002] The research level of apolipoprotein AⅡ is not as good as that of apolipoprotein AⅠ. It is also the second major apolipoprotein in serum high-density lipoprotein, accounting for about 20-25% of the total protein in high-density lipoprotein. It accounts for 15% in HDL2 and 25% in HDL3. Its content in chylomicrons can reach 7-10% of the total apolipoprotein. There is also a small amount of apoAⅡ in very low density lipoprotein. The concentration of apoAⅡ in serum is also higher, second only to apoAⅠ(1.00~1.50g / L) and apoB100(0.8~1.00g / L): 0.35~0.50g / L. In mammals, apoAⅡ is also mainly synthesized by the liver and small intestine. It is a single-chain polypeptide composed of 77 amino acids, often in the form of dimers, with a molecular weight of approximately 8,700da and an isoe...

Claims

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Application Information

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IPC IPC(8): C07K14/775C07K1/16G01N27/447
CPCC07K14/775G01N27/447
Inventor 胡国良曹华斌郭小权张彩英俞海峰
Owner 胡国良
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