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Identifying method of hybridoma cell chromosome

A hybridoma cell and identification method technology, applied in the field of chromosome identification in hybridoma cell subculture, can solve problems such as uneven identification methods and conditions, lack of uniform standards, etc., and achieve the effect of good dispersion and clear pictures

Inactive Publication Date: 2018-11-27
ZHENGZHOU IMMUNO BIOTECH
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Problems solved by technology

[0004] Although there are reports on chromosome identification methods at present, there is a lack of uniform standards, and the identification methods and conditions are uneven, making it difficult to prepare high-resolution chromosome slides with more metaphase divisions and better chromosome dispersion at one time.

Method used

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  • Identifying method of hybridoma cell chromosome

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Embodiment Construction

[0019] The method of the present invention will be described in more detail below through specific examples.

[0020] The identification method of hybridoma cell chromosome of the present invention comprises the following steps:

[0021] In the first step, add colchicine to the hybridoma cells to be tested in a good growth state (covering 70-80% of the bottom of the well) and in the exponential growth phase in a 6-well plate, and adjust the final concentration of colchicine to 0.05μg / ml, 37℃, 5% CO 2 Cultivate in an incubator for 4 hours to stop cell division in metaphase; since colchicine can prevent metaphase cells from continuing to divide, the concentration and time of treatment are very critical. Although high concentration and long-term treatment can obtain more metaphase cells, At the same time, it will also lead to excessive condensation of chromosomes or cleavage of centromere, which will eventually cause abnormal chromosome shape, or even be destroyed or dissolved; ...

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Abstract

The invention discloses an identifying method of hybridoma cell chromosome. The identifying method includes: adding colchicine into to-be-detected hybridoma cells in an exponential growth phase for treatment, collecting cells, adding a hypotonic solution for treatment, using fixing liquid prepared from methanol and glacial acetic acid according to a proportion of 3:1 for first-time prefixing and second-time prefixing, dripping, performing Giemsa dyeing, observing, amplifying, photographing, and counting. The identifying method has the advantages that by limiting parameters and key steps and conditions for making chromosome slide, the obtained chromosome slide is clear in picture and high in dispersity, and number and morphological characteristics of cell chromosome can be observed clearly;genetic stability of the hybridoma cells is judged by analyzing the number and the morphological characteristics of the chromosome, and further studying of the genetic stability of the hybridoma cells through chromosome nucleation analysis technology is facilitated.

Description

technical field [0001] The invention relates to the subculture of hybridoma cells, in particular to a chromosome identification method in the subculture of hybridoma cells. Background technique [0002] Since Kohler and Milstein established lymphocyte hybridoma technology in 1975, scientists have never stopped studying hybridoma cells and monoclonal antibodies. Hybridoma cells have the characteristics of parental cells, which can proliferate indefinitely and be immortal in vitro culture like myeloma cells, and can synthesize and secrete specific antibodies like splenic lymphocytes. The antibody produced by the hybridoma cell line obtained by cloning is a highly homogeneous antibody targeting the same antigenic determinant, the so-called monoclonal antibody (monoclonal antibody), referred to as monoclonal antibody. However, during the subculture of hybridoma cells, the chromosomes encoding antibodies are easily lost, so the identification of hybridoma cell chromosomes is par...

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/68
Inventor 姚营珂鲁盛涛张娟尚朋林赵巧辉李桂林付光宇吴学炜
Owner ZHENGZHOU IMMUNO BIOTECH
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