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Chemically-blocked Antibody Microarray for Multiplexed High-throughput Profiling of Specific Protein Glycosylation in Complex Samples
Published on: May 4, 2012
Chitinase 3-like 2 protein monoclonal antibodies
Stanislav Avdieiev1, Liliia Savinska, Valeriy Filonenko
1Department of Biosynthesis of Nucleic Acids, Institute of Molecular Biology and Genetics, National Academy of Sciences of Ukraine, 150 Zabolotnogo str.Kyiv, Ukraine.
Abstract:
Chitinase 3-like 2 (CHI3L2) is one of the most overexpressed genes in glioblastoma. Despite this, both the CHI3L2 gene and its protein product CHI3L2 are poorly characterized. Here we report the generation and characterization of monoclonal antibodies to CHI3L2 protein (CHI3L2 MAbs). Bacterially expressed 6 His-tagged full-length CHI3L2 was used as antigen. Spleen cells from immunized mice were collected and fused with SP2/0 myeloma cells. Hybridoma clones 2D3 and 4D2 producing high titer CHI3L2 MAbs were identified by enzyme-linked immunosorbent assay (ELISA) and further examined for their activity with the CHI3L2 protein by Western blot analysis and immunoprecipitation. The 2D3 clone was chosen for mouse inoculation and ascites formation. Antibodies derived from the ascitic fluid specifically recognized the recombinant CHI3L2 protein and strongly interacted with CHI3L2 in glioblastoma tissue lysate, as determined by Western blot analysis. The antibodies generated may be useful as a tool in various aspects of CHI3L2 investigation.
Insights
Researchers developed new monoclonal antibodies (MAbs) targeting Chitinase 3-like 2 (CHI3L2), a protein overexpressed in glioblastoma. These CHI3L2 MAbs are validated tools for investigating this key glioblastoma biomarker.
Area of Science:
- Biochemistry
- Immunology
- Oncology
Background:
- Chitinase 3-like 2 (CHI3L2) is significantly overexpressed in glioblastoma, a highly aggressive brain tumor.
- Despite its prevalence, the CHI3L2 gene and its protein product remain poorly understood, hindering therapeutic development.
Purpose of the Study:
- To generate and characterize novel monoclonal antibodies (MAbs) specific to the CHI3L2 protein.
- To validate the utility of these CHI3L2 MAbs as research tools for glioblastoma studies.
Main Methods:
- Bacterially expressed, 6 His-tagged full-length CHI3L2 protein was used as an antigen for immunization.
- Hybridoma technology was employed, involving spleen cell fusion with SP2/0 myeloma cells.
- Selected hybridoma clones (2D3 and 4D2) were screened using ELISA, Western blot, and immunoprecipitation.
Main Results:
- High-titer CHI3L2 MAbs were successfully generated and characterized.
- The 2D3 clone, producing potent CHI3L2 MAbs, was selected for ascites production.
- Western blot analysis confirmed that the generated antibodies specifically recognize recombinant CHI3L2 and interact strongly with CHI3L2 in glioblastoma tissue lysates.
Conclusions:
- Novel monoclonal antibodies targeting CHI3L2 have been successfully developed and validated.
- These CHI3L2 MAbs represent valuable tools for advancing research into glioblastoma biology and diagnostics.
- Further investigation using these antibodies can elucidate CHI3L2's role in glioblastoma progression.
