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Classic case of scFv preparation

source:市场部 data:2021-09-30 views:1023

      Single chain Fv (scFv) is a kind of small molecule genetic engineering antibody, which is composed of VH and VL linked by short peptide of 15-20 amino acids. The complete antibody consists of two heavy chains (H) and two light chains (L). After sequencing, the antibody gene sequence is obtained, and the heavy chain variable region (VH) and light chain variable region (VL) of the antibody are connected to form a recombinant gene through the synthetic linker gene. The antibody expressed by the recombinant gene is called single chain antibody (scFv). Compared with the classical antibody molecules, scFv has the characteristics of small molecular weight (about 1 / 6 of the whole antibody molecule), which can spontaneously fold and form a natural structure with antigen binding characteristics. ScFv has a wide application prospect in antiviral, tumor therapy, autoimmune disease treatment, targeted drug therapy and so on.

      The principle and experimental process of obtaining antibody variable region Fab fragment by hybridoma sequencing have been explained in detail in the last public number article. Recently, Wuhan dai'an Biotechnology Co., Ltd. successfully delivered the scFv and sequence of CD1 * * and CD2 * * to an Institute of Chinese Academy of Sciences. I would like to share with you the service content and experimental results of this project.

Project process

Contract to be signed in January 2021

The gene was synthesized in January 2021, and mammalian cells expressed eukaryotic antigen

Mice were immunized in February 2021

In April 2021, cells were fused to prepare monoclonal antibody cell lines

In June 2021, the customer verified the hybridoma supernatant and selected 5 + 3 positive clones

In July 2021, hybridoma was sequenced and the expression vector was constructed

ScFv protein was expressed and delivered in August 2021

In September 2021, the customer verified the scFv and proved to obtain the scFv sequence with high affinity

The whole process without delay, no repetition, complete in one go, perfect ending! Thank you for your active cooperation. Of course, the most important thing is the customer's evaluation: "great.

Share some highlights of the project:

Antigen preparation

CD1**CD2**

 

Diane biological has experienced technical team, through the advanced mammalian transient expression technology, the preparation of eukaryotic antigen, antigen purity is high, no degradation, can be used for animal immunity.

Detection of mouse antiserum by ELISA

 

 

ELISA was used to detect eukaryotic protein antigen. The titer of mouse antiserum ELISA was more than 1:160000. The high titer could be used for mouse fusion and screening of monoclonal cell lines

Flow cytometry results of monoclonal antibody

 

 

The supernatant of the selected monoclonal cell lines was verified by flow cytometry. The results of customer verification showed that many cell lines achieved the desired results. These cell lines were sequenced to obtain the antibody sequence.

Results of scFv expression

     CD1**scFV    CD2**scFV

 

 

 

The scFv sequence obtained from hybridoma sequencing was used to construct the expression vector and express scFv antibody with high purity.

 ScFv antibody validation results

 

 

 

The results of scFv validation showed that the effect of scFv met the requirements of customers.

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