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Method of quickly measuring titer of ascovirus

A virus titer and rapid determination technology, applied in the direction of biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of insignificant cell lesions, no measurement of vesicle virus titer, etc., to shorten the operation time, shorten the Detection time, the effect of short time consumption

Inactive Publication Date: 2016-10-26
HUAZHONG AGRI UNIV +1
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Problems solved by technology

However, because the cytopathy caused by vesicle virus infecting insect cells is not obvious, the possibility of missed detection and misjudgment is very high, that is, subjective and empirical factors have a greater impact on the detection results, so the traditional endpoint dilution method is used to determine the vesicles. Viral titers are highly flawed
[0006] In summary, due to the special cytopathological characteristics of vesicle virus and the obvious defects of existing virus titer determination methods, there is currently no quick and easy method for measuring vesicle virus titer.

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  • Method of quickly measuring titer of ascovirus
  • Method of quickly measuring titer of ascovirus
  • Method of quickly measuring titer of ascovirus

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Embodiment 1

[0034] The rapid determination of embodiment 1 vesicular virus HvAV-3h titer

[0035] Specific steps are as follows:

[0036] 1. Preparation of gradient dilutions of vesicular virus HvAV-3h

[0037] The vesicle virus liquid HvAV-3h to be tested was diluted with Sf-900II SFM medium, and vortexed for 30 seconds to mix. The specific dilutions are shown in Table 1.

[0038] Table 1 Vesicular virus dilution scheme

[0039]

[0040] 2. Preparation of Insect Cell Suspension

[0041] Take a sufficient amount of Sf9 insect cell culture to collect insect cells according to the conventional subculture method, discard the supernatant medium after standing for 30 minutes, replace with fresh Sf-900II SFM medium, blow with a gun to suspend the insect cells, and count with a hemocytometer The number of cells, and the cells were diluted to a concentration of 1×10 with Sf-900IISFM medium 5 individual / mL.

[0042] 3. Preparation of Cellular Virus Suspension

[0043] Take 1300 μL of insec...

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Abstract

The invention relates to the technical field of biology, and particularly discloses a method of quickly measuring titer of ascovirus. The method includes: resuspending logarithmic phase Sf9 insect cells; placing into an incubator, standing for culture for 30 min until the cells are attached to walls, abandoning a supernatant culture medium, and replacing with a new Sf-900IISFM culture medium; utilizing a blood ball counting plate to measure cell concentration; taking and well mixing Sf9 insect cell suspension with ascovirus diluent, and inoculating to a 96-hole cell culture tray; disposing in the incubator for culture for 24 h, adding trypan blue dyeing liquid into each hole, and recording dyed cell number and total cell number of each hole; calculating Sf9 insect cell death rate of each hole, determining that the corresponding hole is infected with ascovirus if cell death rate of each hole is lower than that of a control group, determining that the corresponding hole is not infected with ascovirus if not, and calculating TCID50 of ascovirus according to a formula. The method combines cell dyeing, can be used for quick measuring of titer of ascovirus and is more accurate in measuring result.

Description

technical field [0001] The invention relates to a method for measuring virus titer, in particular to a method for rapidly measuring vesicle virus titer. Background technique [0002] Ascoviruses are a class of DNA viruses that mainly infect insects of the Lepidoptera family Noctuidae. The vesicular virion contains a circular supercoiled double-stranded DNA, and the viral genome size varies from 116kbp (DpAV4) to 185kbp (HvAV3) depending on the species. The vesicular virus genome contains two sets of repeats. The first group varies in size from 1.1kbp to 3.8kbp according to the species, and is currently known to have no special role in physiology. The second group is about 0.3-1.2kbp in size, encoding the sequence of the baculovirus repeated open reading frame gene. After the host larvae were infected by the vesicle virus, the host larvae showed symptoms such as growth retardation, decreased muscle elasticity, and decreased food intake. Cytopathological features show that...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/06
CPCC12Q1/06
Inventor 万虎韩宁宁李建洪陈子姝黄国华
Owner HUAZHONG AGRI UNIV
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