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Yolk antirotavirus antibody preparation, method for preparing same and use thereof

A rotavirus and preparation technology, applied in the direction of antibody, drug combination, digestive system, etc., can solve the problem of weakening, achieve high neutralizing activity, simple and practical effect

Inactive Publication Date: 2003-08-13
上海海欣生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The protective effect of IgY is related to the time interval between infection with HRV and taking IgY, and it weakens with the increase of the interval time

Method used

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  • Yolk antirotavirus antibody preparation, method for preparing same and use thereof
  • Yolk antirotavirus antibody preparation, method for preparing same and use thereof
  • Yolk antirotavirus antibody preparation, method for preparing same and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1: Isolation of Rotavirus

[0036] Isolation of human pathogenic rotavirus

[0037] Take 10ml of fecal juice from the child with diarrhea, dilute it 1:5 with 0.15M PBS with pH=7.6, put it into a sterile centrifuge tube, shake it fully for 30 minutes, add trypsin with a final concentration of 10μg / ml, and dilute the resulting solution in Incubate at 37°C for 20min. Aspirate the suspension and centrifuge at a high speed of 10000rpm / min. After the supernatant was filtered through a 0.22um filter, the filtrate was serially diluted 10 times with a virus proliferation culture note (MEM medium added with 2 μg / ml trypsin solution), and the resulting solution was used for the next step of amplification and Identification.

[0038] After the Vero cells were cultured to a single layer in a 6-well culture plate, the culture medium was discarded, and 1 ml of each virus isolation material mentioned above was added to each culture well. After incubating at 37°C for 60 min,...

Embodiment 2

[0040] Embodiment 2: In vitro culture and amplification of rotavirus

[0041] The isolated rotavirus culture is mixed with the rotavirus strain which is the standard human type Wa virus strain, and then co-cultivated and amplified in vitro.

[0042] Vero cells (ATCC CCL81) in the logarithmic growth phase were cultured in DMEM-F12 supplemented with 10% FCS (Gibco Company) to form monolayer cells, and the culture area of ​​the culture flask was 150 cm2. Prepare a single cell suspension after digestion with 0.25% trypsin, inoculate into a roller bottle, the initial cell concentration is 1×10 6 / ml, when the cells were cultured to be close to monolayer (80%), the culture medium was replaced with DMEM-F12 culture medium without 10% FCS and added with trypsin to a final concentration of 2 μg / ml. After the mixed virus strain preparation was treated with trypsin 20 μg / ml for 20 minutes, it was inoculated into a roller bottle, and the inoculated virus amount was 2×10 5 For TCID virus...

Embodiment 3

[0043] Embodiment 3: the preparation of rotavirus antigen

[0044] 1. Purification of Virus

[0045] 8L of virus suspension collected after in vitro amplification. Using a Millipore Pellicon ultrafiltration device, the ultrafiltration concentration was 10 times at 4°C, and the molecular weight cut-off was 30KD. The buffer was then switched using 50 mM Tris / HCl, pH=8.0.

[0046] Load the above preparation to a DEAE-Sepharose FF column (2.6×10cm) equilibrated with 50mM Tris / HCl, pH=8.0 in advance, and after fully eluting unbound components, use a linear gradient of 0-0.5M NaCl to elute . Combine virus-containing components for later use.

[0047] 2. Virus inactivation

[0048] The concentrated and purified rotavirus preparation can be inactivated with formalin, phenol, glutaraldehyde, β-propiolactone and other inactivating agents before use. If formalin is used, the dosage between 0.01-0.5%, and the optimal concentration used in the present invention is 0.1%. The inactiva...

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Abstract

An immune globulin composition containing chicken yolk resisting group-A rotaviruses, its preparing process, and its application in treating baby rotavirus diarrhea are disclosed. It is an enteric preparation which is prepared through immunizing hen, biochemical separation and purifying.

Description

technical field [0001] The invention belongs to the field of biotechnology and relates to a novel immunoglobulin, more specifically, the invention relates to an anti-group A rotavirus egg yolk immunoglobulin preparation and its preparation method and application. It is suitable for the prevention and treatment of intestinal diseases such as diarrhea in infants and young children. Background technique [0002] Infant diarrhea is a common disease in infants and young children, and it accounts for the second place in the incidence of all diseases in emergency cases and hospitalized cases (after respiratory infections). 40 million, because the physiological development of infants is not complete, once they get sick, many serious complications often occur, and the mortality rate is relatively high. According to statistics, the number of infants and young children who die from diarrhea is more than 8 million every year in the world, especially in developing countries, because the...

Claims

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Application Information

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IPC IPC(8): A61K39/395A61P1/12
Inventor 王建莉张立煌王青青
Owner 上海海欣生物技术有限公司
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