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A device and method for inactivating rabies virus and hydrolyzing an inactivating agent using a disposable bag

A rabies virus and virus inactivation technology, which is applied in the direction of viruses, virus/bacteriophage, biochemical cleaning equipment, etc., can solve the problems of antigen loss and insufficient inactivation, and achieve low risk of damage, reduce material splash, reduce The effect of antigen loss

Active Publication Date: 2019-03-05
SHANDONG YIDU BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Using a 20L barrel or stainless steel tank requires magnetic stirring or a shaker to mix the inactivator and the virus stock solution evenly. During the mixing process, a large amount of foam will be generated due to vigorous shaking, which may easily cause insufficient inactivation and loss of antigens.

Method used

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  • A device and method for inactivating rabies virus and hydrolyzing an inactivating agent using a disposable bag
  • A device and method for inactivating rabies virus and hydrolyzing an inactivating agent using a disposable bag

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] The large-scale inactivation method of embodiment 1 rabies vaccine virus liquid:

[0041] Take the prepared Vero cell rabies virus harvest liquid and filter it through 0.8μm and 0.45μm microporous filters. After clarification and filtration, use Millipore's 300KD ultrafiltration membrane bag for ultrafiltration. The washing liquid (BME liquid, containing 0.02% glutamine by mass percentage, 24.63% sodium bicarbonate) is used to balance the membrane package, and the pH value of the reflux liquid is detected, and the pH value is 6.0. After meeting the requirements, it is concentrated, and the concentration ratio is 25 times. Transfer the virus pool from port A to the inactivated disposable bag 10. First dilute β-propiolactone with sterile water for injection 1:40 times, and then add it to the concentrated virus solution from port B of the bag according to the volume ratio of 1:100, and use PBS solution to wash from port B. Close the EZD valve A8, place in a cold storage a...

Embodiment 2

[0042] Embodiment 2 Inactivator hydrolysis situation:

[0043] The hydrolysis of β-propiolactone at 37°C was monitored by gas chromatography analysis method. Analysis of hydrolysis of β-propiolactone. Through the analysis of the test results, the concentration of β-propiolactone dropped from 299.4ppm when it was not hydrolyzed to 34.4ppm when it was hydrolyzed for 150min, which was lower than the quantitative limit of the method, indicating that β-propiolactone could be completely hydrolyzed at 37°C for 150min. hydrolysis effect.

Embodiment 3

[0044] Embodiment 3 cell method verification inactivation effect:

[0045] According to the inactivation method of the embodiment, take the virus stock solution according to every 3cm 2 Inoculate the inactivated rabies virus liquid into MRC-5 cells at a ratio of 1ml for cell inoculation, culture for 7 days and then subculture, continue to culture for 21 days, inoculate the cell suspension on Lab-tek chamber slides (purchased from Thermo Company), After 3-4 days, Lab-tek was observed for the presence of rabies virus by immunofluorescence. Among them, the cell maintenance solution (MEM solution containing 4% fetal bovine serum) was replaced on the 14th day and 21st day respectively, and the cell supernatant was collected for mouse brain cavity poisoning, and the mice were observed for 21 days. A cell negative control and a virus positive control were also set up in the experiment to determine the validity of the experiment.

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Abstract

The invention provides a device and method for rabies virus inactivation and inactivator hydrolysis by virtue of disposable bags, wherein an inactivating disposable bag is provided with three connecting ports; the three connecting ports include an inflating port which is arranged at the upper side and is connected to an inflating group, and a liquid inlet and a liquid outlet which are arranged atthe bottom; the liquid inlet is connected to a liquid filling group and the liquid outlet is connected to a drainage group; a built-in stirrer is arranged in the center of the bag, wherein the inflating group is composed of a bag-type filter, a silicone tube and a pipeline clamp, and the sterile bag covers the external part of the bag-type filter; the liquid filling group is formed by sequentiallyconnecting a connector B, a silicone tube, a pipeline clamp, a tee joint and an EZD valve; and another opening of the tee joint is connected to a silicone tube, a pipeline clamp and a connector A. The device provided by the invention has the advantages that the disposable bags are transformed and reasonably used, so that a virus liquid inactivation / hydrolysis method is improved; therefore, a riskof failing in virus inactivation is reduced, antigen loss is lowered and production cost is reduced.

Description

technical field [0001] The invention relates to the field of virus vaccine production, in particular to a device and method for inactivating rabies virus and hydrolyzing an inactivating agent using a disposable bag. Background technique [0002] The containers used for the inactivation of the virus stock solution in the current vaccine production workshop are mainly 20L Nalgene plastic buckets or stainless steel tanks. Disposable bags are mainly used for the preparation, storage and transfer of solutions required in production. Before using 20L barrels or stainless steel tanks for rabies virus inactivation, autoclaving, cleaning verification, and detection of detergent / disinfectant residues are required. Disposable bags have complete manufacturer verification support and can be used directly for large-scale production It can reduce the cleaning and sterilization links, reduce the space occupied by equipment, and the cost is relatively low. [0003] Using a 20L bucket or st...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12M1/00C12M1/04C12M1/02C12N7/00C12N7/06
CPCC12M21/10C12M23/14C12M27/02C12M29/06C12M41/22C12N7/00C12N2760/20151C12N2760/20163
Inventor 庄长鹰
Owner SHANDONG YIDU BIOTECH
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