Abstract
Objective:
To explore the adjuvant effects of Mycobacterium tuberculosis heat shock protein (TBhsp) and Mycobacterium tuberculosis T cell stimulatory epitope (MT) in the preparation of the monoclonal antibody (mAb) against phosphatase of regenerating liver-3 (PRL-3).
Methods:
The prokaryotic expression vectors pET28a were used for the construction of pET28a-PRL-3 (control plasmids), pET28a-PRL-3-MT, pET28a-TBhsp-PRL-3 and pET28a- TBhsp-PRL-3-MT (recombinant plasmids). The various fusion proteins expressed in bacteria were purified and utilized to immunize the BALB/c mice, respectively. The serum level of specific anti-PRL-3 antibody was measured by ELISA. The mouse with the highest serum level of anti-PRL-3 antibody was selected to prepare mAb with hybridoma technique. The subclasses of mAb were identified.
Results:
The PRL-3 fusion proteins expressed from the four expression plasmids were purified successfully. The serum level of anti-PRL-3 antibody from the mice immunized with PRL-3-MT was the highest compared with the other immune groups. Ten hybridoma cell lines secreting anti-PRL-3 mAb were obtained after cell fusion and ELISA primary screening, and all of the mAb subclasses were IgG.
Conclusion:
MT is a potentially effective molecular adjuvant in preparation of mAb specific for PRL-3.
MeSH terms
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Adjuvants, Immunologic / genetics
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Adjuvants, Immunologic / metabolism
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Amino Acid Sequence
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Animals
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Antibodies, Monoclonal / blood
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Antibodies, Monoclonal / immunology*
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Antibody Specificity / immunology
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Bacterial Proteins / genetics
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Bacterial Proteins / immunology*
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Bacterial Proteins / metabolism
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Electrophoresis, Polyacrylamide Gel
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Enzyme-Linked Immunosorbent Assay
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Epitopes, T-Lymphocyte / genetics
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Epitopes, T-Lymphocyte / immunology*
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Epitopes, T-Lymphocyte / metabolism
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Female
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Heat-Shock Proteins / genetics
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Heat-Shock Proteins / immunology*
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Heat-Shock Proteins / metabolism
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Immediate-Early Proteins / genetics
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Immediate-Early Proteins / immunology*
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Immediate-Early Proteins / metabolism
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Immunization
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Immunoglobulin G / blood
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Immunoglobulin G / immunology
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Mice
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Mice, Inbred BALB C
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Molecular Sequence Data
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Mycobacterium tuberculosis / genetics
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Mycobacterium tuberculosis / metabolism
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Protein Tyrosine Phosphatases / genetics
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Protein Tyrosine Phosphatases / immunology*
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Protein Tyrosine Phosphatases / metabolism
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / immunology*
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Recombinant Fusion Proteins / metabolism
Substances
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Adjuvants, Immunologic
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Antibodies, Monoclonal
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Bacterial Proteins
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Epitopes, T-Lymphocyte
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Heat-Shock Proteins
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Immediate-Early Proteins
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Immunoglobulin G
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Ptp4a3 protein, mouse
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Recombinant Fusion Proteins
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Protein Tyrosine Phosphatases