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Preparation method of swine leukocyte interferon

A leukocyte interferon and leukocyte technology, applied in the field of biological preparations, can solve the problems of high production cost of porcine leukocyte interferon, affecting the application of porcine leukocyte interferon, low leukocyte activity and other problems, and achieves low cost, stable and reliable performance, leukocyte The effect of improving the purity

Inactive Publication Date: 2014-10-01
张永奎
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

As a biological product, PLeIFN not only treats directly, but also is safe and effective. At the same time, the use of PLeIFN has important clinical significance for porcine viral diseases that currently have no effective drug treatment.
However, in the existing PleIFN preparation process, due to reasons such as low leukocyte activity and excessive foreign protein, the production cost of the existing porcine leukocyte interferon is too high, the product quality is difficult to control, and the clinical curative effect is unstable, which seriously affects the Application of Porcine Leukocyte Interferon

Method used

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  • Preparation method of swine leukocyte interferon
  • Preparation method of swine leukocyte interferon
  • Preparation method of swine leukocyte interferon

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1, such as figure 1 Shown, a kind of preparation method of porcine leukocyte interferon comprises the following steps:

[0042] Step 1), add the fresh anticoagulated blood collected by aseptic method into an equal volume of 30g / L gelatin saline, mix well and put it in a 37°C incubator until the layers are clear, take the supernatant and centrifuge at 3000rpm After 15 minutes, discard the supernatant and resuspend the pellet;

[0043] Step 2) Take the precipitate collected in step 1 and add ammonium chloride solution with a mass volume concentration of 0.83% according to the volume ratio of 1:1, mix well and then centrifuge at 3000rpm for 15min, take the precipitate after the first centrifugation again according to the ratio of 1:1 Add 0.83% ammonium chloride solution, mix well and centrifuge at 3000rpm for 15min, wash the precipitate after the second centrifugation with Hanks solution three times, add 1 to 1.5 times the volume of the original whole blood in Ea...

Embodiment 2

[0056] Embodiment 2, a kind of preparation method of porcine leukocyte interferon, comprises the following steps:

[0057] Step 1), add the fresh anticoagulated blood collected by aseptic method into an equal volume of 30g / L gelatin saline, mix well and put it in a 37°C incubator until the layers are clear, take the supernatant and centrifuge at 3000rpm After 15 minutes, discard the supernatant and resuspend the pellet;

[0058] Step 2) Take the precipitate collected in step 1 and add ammonium chloride solution with a mass volume concentration of 0.83% according to the volume ratio of 1:1, mix well and then centrifuge at 3000rpm for 15min, take the precipitate after the first centrifugation again according to the ratio of 1:1 Add 0.83% ammonium chloride solution, mix well and centrifuge at 3000rpm for 15min, wash the precipitate after the second centrifugation with Hanks solution three times, add 1 to 1.5 times the volume of the original whole blood in Eagle's with 5% porcine ...

Embodiment 3

[0090] Example 3, the porcine leukocyte interferon prepared by the above preparation method has the following research on Haemophilus parasuis disease.

[0091] Clinical symptoms:

[0092] Haemophilus parasuis: including fever, dyspnea, joint swelling, lameness, cyanosis of skin and mucous membranes, difficulty standing or even paralysis, stiff pigs or death. Sows may suffer from miscarriage, and boars may have lameness. Lameness in lactating sows can lead to an extreme weakening of motherhood. When he died, his body surface was purple, his stomach was big, there was a large amount of yellow ascites, and a large amount of cellulose exuded from the mesentery, especially the liver was completely encased, and the interstitial edema of the lungs was swollen.

[0093] Efficacy evaluation criteria:

[0094] Cure: After injecting the porcine leukocyte interferon of the present invention, the disease symptoms completely disappear, and the main physical and chemical examination inde...

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Abstract

The invention belongs to the technical field of biological agents, and particularly relates to a preparation method of swine leukocyte interferon. The preparation method comprises the steps of adding gelatin normal saline for layering, centrifugation and heavy suspension precipitation, adding an ammonium chloride solution for centrifugation and culture, adding a Newcastle disease virus NDV II for rotary culture, adjusting a PH (potential of hydrogen) value with NaOH and Hcl, centrifuging to remove a precipitate to form a swine leukocyte interferon crude solution, adding the swine leukocyte interferon crude solution into sterile potassium thiocyanate for dissolution, then carrying out acidizing, adding the centrifuged precipitate into an alcohol solution by an atomization method, centrifuging to remove a precipitate, adjusting a PH value, then sequentially adding NaAc and KCNS for centrifugal precipitation, adding a precipitate into a PBS (phosphate buffer solution) for dissolution and dialysis, and obtaining the refined swine leukocyte interferon. The prepared swine leukocyte interferon has higher activity; the purity of leucocyte is improved greatly by cracking of the ammonium chloride solution; a purification effect of the interferon is better via NaAc; the activity recovery rate is higher; the whole yield of the interferon is increased; and the production cost of the interferon is lowered.

Description

technical field [0001] The invention belongs to the technical field of biological preparations, in particular to a preparation method of porcine leukocyte interferon which can effectively improve the viability and purity of leukocytes. Background technique [0002] Interferon (IFN) is a broad-spectrum antiviral agent. It does not directly kill or inhibit the virus, but mainly through the action of cell surface receptors to make cells produce antiviral proteins, thereby inhibiting the replication of hepatitis B virus. Its types are divided into three types type, α-(leukocyte) type, β-(fibroblast) type and γ-(lymphocyte) type; at the same time, it can also enhance the activity of natural killer cells (NK cells), macrophages and T lymphocytes, thereby To the immune regulation, and enhance the antiviral ability. Interferon was first discovered by Issacs and Lindeman in 1957. It is a group of active proteins (mainly glycoproteins) with multiple functions and a cytokine produced ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/555C07K1/30
CPCC07K14/555
Inventor 李勉张永奎郭婧贾丽敏
Owner 张永奎