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Preparation method of SETD3 antibody for immunofluorescence assay

An immunofluorescence detection and antibody technology, applied in the field of immunology, can solve the problem of low titer and achieve the effect of high titer and specificity

Inactive Publication Date: 2017-06-27
SHENZHEN RES INST OF WUHAN UNIVERISTY
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the titer of Atlas antibody is not high, and the dilution ratio is greater than 1:1000

Method used

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  • Preparation method of SETD3 antibody for immunofluorescence assay
  • Preparation method of SETD3 antibody for immunofluorescence assay

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Embodiment Construction

[0027] In order to fully understand the technical content of the present invention, the technical solution of the present invention will be further introduced and illustrated below in conjunction with specific embodiments.

[0028] A preferred embodiment of the preparation method of SETD3 antibody for immunofluorescence detection:

[0029] 1. Clone the full-length human SETD3 protein gene with N-terminal 6xHis and GST tags, and express it in E. coli.

[0030] 2. Purify N-terminal 6xHis-tagged SETD3 protein with nickel column resin, and 5 mg of GST-tagged SETD3 protein with glutathione resin.

[0031] 3. Simultaneously immunize two New Zealand big rabbits with purified N-terminal His-tagged SETD3 protein, main injection of 0.5mg N-terminal His-tagged SETD3 protein antigen, 0.6mg N-terminal His-tagged SETD3 protein antigen on the 28th day after the main injection, and after the main injection Inject 0.6 mg N-terminal His-tagged SETD3 protein antigen on the 42nd day, inject 0.4 ...

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Abstract

The invention discloses a preparation method of an SETD3 antibody for immunofluorescence assay. The method comprises the following steps: immunizing rabbit with protein expressed through full-length SETD3 which is labelled with N-terminal 6xHis, and extracting serum of the immunized rabbit; and purifying the serum of the rabbit with the full-length SETD3 protein which is labelled through GST that is coupled with Affi-Gel. The SETD3 antibody is applicable to western blot (WB), immunofluorescence (IF), immunohistochemistry (IHC) and immunoprecipitation (IP), and is high in titer and specificity.

Description

technical field [0001] The invention relates to the technical field of immunology, in particular to a method for preparing a SETD3 antibody used for immunofluorescence detection. Background technique [0002] SETD3 is a methyltransferase with a classic SET conserved structure (Su(var)3-9, Enhancer-of-zeste and Trithorax). SETD3 itself also contains a domain called Rubis-subs-bind. SETD3 was first identified because its transcript levels are regulated during feeding. Studies have found that SETD3 in zebrafish has the function of single, double and trimethylation of the 36 residues of histone H3K and the function of inhibiting cell viability by inducing the activation of caspase-3. Recent studies have shown that mouse-derived SETD3 can induce the differentiation process of muscle cells by binding to the promoter of myogenic factor myogenin, so mouse-derived SETD3 plays the role of transcription factor here, correspondingly, mouse-derived SETD3 was shown to have methyltransf...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/40C07K16/06G01N33/531
CPCC07K16/40C07K16/06C07K16/065G01N33/531
Inventor 杜海宁程晓庆赵梦洁
Owner SHENZHEN RES INST OF WUHAN UNIVERISTY
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