Stabilized denatured lipoprotein and process for producing the same

A lipoprotein and stabilization technology, applied in the field of preparation of denatured lipoprotein, stabilized denatured lipoprotein and its preparation, can solve the problems of unsatisfactory storage stability and cumbersome operation of standard substances

Inactive Publication Date: 2002-09-04
KYOWA MEDEX CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, since the storage stability of the standard substance mentioned in the above method is not very ideal, it must be re-prepared every time it is used immediately, so the operation appears to be loaded down with trivial details

Method used

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  • Stabilized denatured lipoprotein and process for producing the same
  • Stabilized denatured lipoprotein and process for producing the same
  • Stabilized denatured lipoprotein and process for producing the same

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Experimental program
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no. 4 approach

[0057] According to the fourth embodiment of the present invention, the present invention provides the stabilized denatured lipoprotein prepared by the above-mentioned first or third embodiment.

[0058] The denatured lipoprotein and the stabilized denatured lipoprotein prepared by the above steps have excellent long-term storage stability, for example, they are brought into contact with an antibody that recognizes the denatured lipoprotein, and the reactivity of the antibody to the sample is measured to measure It can be used as a standard substance in methods containing denatured lipoproteins, and can also be used as various experimental reagents for studying the physiological functions and activities of denatured lipoproteins.

no. 5 approach

[0059] Therefore, according to the fifth embodiment of the present invention, the present invention provides a method for measuring denatured lipoprotein, which method uses the stabilized denatured lipoprotein prepared by the above-mentioned first or third embodiment as a standard substance. In addition, according to the sixth embodiment of the present invention, the present invention provides a kit for measuring denatured lipoprotein, which kit contains the stabilized denatured lipoprotein prepared by the above-mentioned first or third embodiment as a standard substance.

[0060] In the above-mentioned fifth embodiment, the method of measuring denatured lipoprotein is not particularly limited, and a known method can be used. Specifically, denatured lipoproteins are brought into contact with antibodies that recognize denatured lipoproteins, and the reactivity of the antibodies to a sample is measured to measure denatured lipoproteins contained in blood components. For example, ...

Embodiment 1

[0065] (1) Preparation of LDL

[0066]Obtain ultracentrifugation from human plasma obtained with EDTA as an anticoagulant (10°C, adjust the specific gravity to 1.019, 120000×g, centrifuge for 20 hours, recover the upper layer, adjust the specific gravity to 1.063, and centrifuge at 120000×g for 24 hours) The portion with a specific gravity of 1.019 to 1.063 is recovered as an LDL component. At this time, the purity of LDL was confirmed by obtaining a single sharp band in agarose gel electrophoresis.

[0067] Next, the LDL fraction was purified by sufficient dialysis (16 hours or overnight) against PBS (pH 7.4) containing 0.25 mmol / L EDTA. The above-mentioned purified LDL was then dissolved in PBS (pH 7.4) containing 0.25 mmol / L EDTA to make the LDL protein concentration 1 mg / mL, and stored at 4°C for future use. In this example, the protein content was determined by Lowry's improved method. Briefly, 2(w / v)% sodium carbonate, 0.4(w / v)% sodium hydroxide, 0.16(w / v)% tartaric a...

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Abstract

Denatured lipoprotein excelling in stability of prolonged preservation (namely exhibiting only sparing liability to induce denaturation of lipoprotein) and used as a standard substance for the determination of denatured lipoprotein in blood or the determination of the physiological activity of the lipoprotein and a method for the production thereof are provided. The method for the production of the stabilized denatured lipoprotein comprises artificially denaturing lipoprotein, thereby obtaining denatured lipoprotein, freeze-drying the denatured lipoprotein or performing a process including at least one freezing operation on a solution containing denatured lipoprotein thereby denaturing the lipoprotein contained in the solution and obtaining denatured lipoprotein and further freeze-drying the denatured lipoprotein.

Description

technical field [0001] The invention relates to a stabilized denatured lipoprotein and a preparation method thereof. Specifically, the present invention relates to a denatured lipoprotein having excellent long-term storage stability obtained by denaturing the lipoprotein and then freeze-drying the denatured lipoprotein and a preparation method thereof. [0002] The invention also relates to the preparation method of denatured lipoprotein. Specifically, the method for preparing denatured lipoproteins involved in the present invention is a method for denaturing lipoproteins by subjecting a solution containing lipoproteins to a step including at least one freezing step, and the present invention also relates to reproducing the denatured lipoproteins thus prepared. Denatured lipoprotein with excellent long-term storage stability obtained by freeze-drying and a preparation method thereof. [0003] Denatured lipoproteins are known to be associated with diseases of the coronary art...

Claims

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Application Information

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IPC IPC(8): C07K14/775
CPCC07K14/775
Inventor 重松贵岛村京子木村顺治河野弘明末重信之
Owner KYOWA MEDEX CO LTD
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