Method for detecting titre of live viruses

Abstract

The invention discloses a method for detecting the titre of live viruses, which comprises the following steps: firstly, infecting diploid cells or other adherent cells with dilute viruses to be detected, culturing and fixing the infected cells; secondly, adding virus antiserum (first antibody) into the fixed cells, culturing and washing, adding a second antibody marked by horseradish peroxidase (HRP), culturing and washing, adding coloring solution for color development and producing red or brown spots, or adding virus antiserum marked by the HRP into the fixed cells, culturing the virus antiserum and washing, adding the coloring solution for color development and producing red or brown spots; thirdly, calculating the virus titre according to the quantity of the red or brown spots and the dilution multiple, or calculating the 1gTCID50 value of the viruses according to the dilution degree and the hole numbers of the spots. The detection method can be applicable to the titre detection of various live viruses, can shorten the detection time of the titre of the viruses due to fast detection speed, has good specificity as the immune spots are caused by special immune reaction, low detection cost as well as simple and convenient operation and does not need special instruments of optical or fluorescent microscopes, and the like or other reagents.

Description

technical field [0001] The invention relates to a method for detecting the titer of a live virus, in particular to a method for detecting the titer of a live virus by using an enzyme-linked immunospot method, and belongs to the field of biopharmaceuticals. Background technique [0002] So far, the detection methods for live virus titer mainly include plaque method, endpoint dilution method and enzyme-linked immunofluorescence method. The main detection process and advantages and disadvantages of the three detection methods are as follows: [0003] Plaque method: This method is the most classic method for determining virus titers. Generally speaking, one plaque is produced by one virus particle, expressed in plaque forming units (PFU). Taking varicella virus as an example, 2BS or MRC-5 cells in good growth state in the culture flask are subcultured to 6-well cell culture plates, 37°C, 5% CO 2 Culture in an incubator until the cells form a monolayer for the experiment. Disca...

AI Technical Summary

Problems solved by technology

There are several defects in this method: 1. The detection time is long, which generally takes 8-10 days; 2. The accuracy is not high. When the virus-infected cells are cultured for 8-10 days, adjacent plaques may unite together, reducing the If the virus titer is lowered, it is also possible that the mother plaque virus produces daughter plaques, which increases the virus titer, which leads to poor repeatability of the test; 3. The number of plaques is only the number, and there is no specificity, so it cannot reflect whether the plaques are infected by varicella virus. caused by disease

The advantage of this method is that the time required for detection is short and it can reflect the specificity of the virus. However, a fluorescent microscope is required, and the operator needs professional experience, and the measured results of different operators vary greatly.

[0006] In short, there is still a lack of a rapid, accurate, specific, and low-cost detection method for the detection of live virus titers.

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