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Method for in vitro induction of specific cytotoxic T lymphocytes of hepatitis B virus (HBV)

A hepatitis B virus, lymphocyte technology, applied in the field of cell biology and immunology of biotechnology, can solve problems such as the inability to completely remove the virus

Inactive Publication Date: 2011-08-31
ZHEJIANG UNIV
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  • Summary
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  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0002] The treatment of chronic hepatitis B virus infection is still a major problem in the medical field. Currently, anti-HBV drugs cannot completely eliminate the virus. How to effectively eliminate HBV is still a hot spot in the treatment of hepatitis B.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0028] 1) Preparation of artificial antigen-presenting cell aAPC that can load any polypeptide:

[0029] Use MHC I-Ig and anti-CD28 mAb to coat magnetic beads: MHC I-Ig and anti-CD28 mAb are coated on magnetic beads (Dynabeads M-450, purchased from invitrogen) by direct coating method, and constructed to load any polypeptide artificial antigen-presenting cell aAPC;

[0030] Specific method: sterilized 0.1M borate buffer washes magnetic beads twice, makes magnetic beads mixed with 1:1 HLA-A2-Ig and anti-CD28 mAb9.3 (monoclonal antibody against CD28, purchased) Incubate at 4°C for 24h on a rotator. Wash twice with bead washing buffer, and then incubate for 24h in buffer containing magnetic beads, then remove the washing solution and add fresh buffer. The prepared aAPC finally contains 0.9×10 per magnetic bead 5 Molecular HLA-A2-Ig and 1.9 x 10 5 anti-CD28 mAb 9.3 molecules. The prepared magnetic beads can be stored at 4°C for 3 months. The coated magnetic beads are aAPC. ...

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PUM

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Abstract

The invention discloses a method for in vitro induction of specific cytotoxic T lymphocytes of hepatitis B virus (HBV). The method comprises the following steps of: 1, preparing artificial antigen presenting cells (aAPC) capable of loading any polypeptides; 2, loading aAPC on epitope peptides; and 3, loading aAPC in vitro induction HBV antigen specific CTL of antigen peptides; measuring the number of obtained antigen specific CTL by a Tetramer decoration method, namely combining streptavidin marked by fluorescein and four MHC-peptide compounds marked by biotin to form the Tetramer, namely MHC-peptide tetramer; and after the MHC-peptide tetramer is combined with TCR on the antigen specific CTL, measuring by a flow cytometry. The method can be applied to the amplification of T lymphocytes resisting the HBV.

Description

technical field [0001] The invention belongs to the fields of cell biology and immunology of biotechnology, and relates to the in vitro treatment of artificial antigen-presenting cells (aAPC) with HBV cytotoxic T lymphocyte (Cytotoxic T Lymphocyte, CTL) epitope peptide Induces HBV-specific CTLs for killing cells expressing HBV antigens. Background technique [0002] The treatment of chronic hepatitis B virus infection is still a major problem in the medical field. The current anti-HBV treatment drugs cannot completely eliminate the virus. How to effectively eliminate HBV is still a hot spot in the treatment of hepatitis B. [0003] (1) HBV-specific cytotoxic T lymphocytes (Cytotoxic T Lymphocytes, CTLs) are an important factor for the body to clear HBV. It is of great significance for the treatment of chronic hepatitis B to study the preparation method of specific CTL with polyclonal, multispecific and strong response. [0004] Studies have shown that HBV-specific CTL is a...

Claims

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Application Information

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IPC IPC(8): C12N5/0783
Inventor 朱海红
Owner ZHEJIANG UNIV
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