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Antibodies recognizing both IgM isotypes in Atlantic salmon
Ida Aagård Hedfors1, Hege Bakke, Karsten Skjødt
1CEES, Department of Biology, University of Oslo, Oslo, Norway. ida.hedfors@bio.uio.no
Fish & Shellfish Immunology
|September 27, 2012
Summary
New monoclonal antibodies (mAbs) effectively identify IgM(+) B cells in Atlantic salmon, enabling better study of fish immune responses. These tools distinguish between IgM-A and IgM-B isotypes, crucial for understanding fish immunology.
Area of Science:
- Immunology
- Fish Biology
- Cell Biology
Background:
- Understanding immune cell subsets is vital for disease defense, yet poorly defined in teleost fish due to limited tools.
- Immunoglobulins (Ig) serve as cell surface markers for immune cell identification, requiring specific antibodies.
- Distinguishing B cell populations in Atlantic salmon (Salmo salar) requires reliable markers for IgM (Immunoglobulin M).
Purpose of the Study:
- To identify and characterize tools for separating IgM-positive (IgM(+)) B cells from IgM-negative (IgM(-)) non-B cells in Atlantic salmon using flow cytometry.
- To evaluate the efficacy of existing and novel monoclonal antibodies (mAbs) and polyclonal antibodies (pAb) against Atlantic salmon IgM.
Main Methods:
- Isolation of leukocytes from peripheral blood, spleen, and head kidney of Atlantic salmon.
- Staining of isolated cells with various commercially available and locally produced mAbs and pAbs targeting salmon IgM.
- Intracellular staining of 293T cells transfected with Atlantic salmon IgM-A and IgM-B constructs to determine antibody isotype specificity.
Main Results:
- Significant variability was observed in staining patterns among tested antibodies, with positively stained cells ranging from <5% to >80%.
- Previously used mAbs (4c10, N2, 1.14) recognized only a fraction of salmon B cells, with 4c10 binding exclusively to the IgM-A isotype.
- Three novel mAbs (IgF1-3, -18, -19) and potentially one commercial antibody (IgH FITC from Cedarlane) demonstrated binding to both IgM-A and IgM-B isotypes.
Conclusions:
- The novel mAbs IgF1-3, -18, and -19 provide effective tools for the complete depletion or enrichment of IgM(+) B cells and/or IgM(-) T cells in Atlantic salmon.
- These new reagents significantly advance the ability to study B cell populations and immune responses in salmonids.
- Accurate identification of IgM-expressing cells is now feasible, facilitating deeper research into fish immunology.
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