Fusion protein TT-B7-H4IgV as well as preparation method and application thereof

A fusion protein, B7-H4 technology, applied in the fusion protein inducing anti-B7-H4 antiserum, T-B7-H4IgV and its preparation field, can solve the problem of limited application of tumor vaccines, lack of antigen presentation, lack of specificity tumor antigens, etc.

Inactive Publication Date: 2009-09-30
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, due to the lack of specific tumor antigens and lack of effective antigen presentation, the current application of tumor vaccines is limited.

Method used

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  • Fusion protein TT-B7-H4IgV as well as preparation method and application thereof
  • Fusion protein TT-B7-H4IgV as well as preparation method and application thereof
  • Fusion protein TT-B7-H4IgV as well as preparation method and application thereof

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Embodiment Construction

[0081] The technical solution of the present invention will be described in detail below in conjunction with the accompanying drawings.

[0082] 1. Construction of recombinant plasmid pQE-30-TT-B7-H4IgV

[0083] Query the human B7-H4 cDNA gene sequence (NM024626) in GeneBank, intercept the IgV sequence of the extracellular segment of B7-H4, and introduce BamHI restriction site and T helper cell epitope upstream, (T helper cell epitope is tetanus toxoid epitope TT830-843, its amino acid sequence is: NSKFIGITE, to trigger a strong cellular immune response in the body, thereby promoting the tumor vaccine to play a better role), the downstream introduced SalI restriction site, and its recombinant sequence is as follows (SEQ.ID. NO.2):

[0084] atgagaggat cgcatcacca tcaccatcac ggatcccagt atataaaagc aaattctaaa 60

[0085] tttataggta taactgaagc agcagcagaa ttcctgagct gcacttttga acctgacatc 120

[0086] aaactttctg atatcgtgat acaatggctg aaggaaggtg ttttaggctt ggtccatgag 180

[0087] t...

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Abstract

The invention relates to a fusion protein TT-B7-H4IgV as well as a preparation method and an application thereof. The B7-H4IgV is a variable region IgV of a B7-H4 molecule extracellular region, TT is a T auxiliary cell epitope peptide or a tetanus toxoid epitope, the N end of the fusion protein TT-B7-H4IgV is connected with 6 His residues, a connection mode is as follows: 6*His-TT-B7-H4IgV, and the fusion protein TT-B7-H4IgV is applied to prepare anti-tumor vaccines which take the B7-H4 as a target spot. The fusion protein TT-B7-H4IgV is established by selecting the B7-H4 as a target spot and taking the TT as antigen presentation and is in accordance with prospects by being identified by a Western blot and checking orders by the Western blot; a purified fusion protein TT-hB7-H4IgV is used for immunizing small Kunming mice, and the biological activity of antiserums is measured by ELISA and flow cytometry so as to prove that a B7-H4 antiserum and the B7-H4 have good combination activity, thus the purified fusion protein TT-hB7-H4IgV can induce organisms to generate anti-B7-H4 polyclonal antibodies.

Description

technical field [0001] The invention belongs to the field of medical biotechnology, and specifically relates to the construction of fusion protein recombinant plasmids, expression in prokaryotic cells, and purification of target proteins, in particular to a fusion protein TT-B7-H4IgV capable of inducing anti-B7-H4 antiserum And its preparation method and use. Background technique [0002] 1. B7-H4 molecule and its research [0003] B7-H4 (B7-S1, B7x) is a new member of the B7 family discovered in 2003 by Chen and other three laboratories using bioinformatics methods. They used the extracellular segment variable region (IgV+IgC) of the existing B7 family members as the query sequence, found it when searching the EST sequence (expressed sequence tag) in GenBank, and obtained it from the human placenta cDNA library full-length sequence. B7-H4 cannot bind to CD28, CTLA-4 and ICOS, it binds to unknown receptors and regulates the immune response. In recent years, more and more...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/70C07K1/22A61K38/16A61P35/00
Inventor 张英起朱文华
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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