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Home / Equine Research Bank / Cytometry A / In search for cross-reactivity to immunophenotype equine mes...
Cytometry. Part A : the journal of the International Society for Analytical Cytology2012; 81(4); 312-323; doi: 10.1002/cyto.a.22026

In search for cross-reactivity to immunophenotype equine mesenchymal stromal cells by multicolor flow cytometry.

Abstract: During recent years, cell-based therapies using mesenchymal stem cells (MSC) are reported in equine veterinary medicine with increasing frequency. In most cases, the isolation and in vitro differentiation of equine MSC are described, but their proper immunophenotypic characterization is rarely performed. The lack of a single marker specific for MSC and the limited availability of monoclonal antibodies (mAbs) for equine MSC in particular, strongly hamper this research. In this study, 30 commercial mAbs were screened with flow cytometry for recognizing equine epitopes using the appropriate positive controls to confirm their specificity. Cross-reactivity was found and confirmed by confocal microscopy for CD45, CD73, CD79α, CD90, CD105, MHC-II, a monocyte marker, and two clones tested for CD29 and CD44. Unfortunately, none of the evaluated CD34 clones recognized the equine epitopes on positive control endothelial cells. Subsequently, umbilical cord blood-derived undifferentiated equine MSC of the fourth passage of six horses were characterized using multicolor flow cytometry based on the selected nine-marker panel of both cell surface antigens and intracytoplasmatic proteins. In addition, appropriate positive and negative controls were included, and the viable single cell population was analyzed by excluding dead cells using 7-aminoactinomycin D. Isolated equine MSC of the fourth passage were found to be CD29, CD44, CD90 positive and CD45, CD79α, MHC-II, and a monocyte marker negative. A variable expression was found for CD73 and CD105. Successful differentiation towards the osteogenic, chondrogenic, and adipogenic lineage was used as additional validation. We suggest that this selected nine-marker panel can be used for the adequate immunophenotyping of equine MSC.
Copyright © 2012 International Society for Advancement of Cytometry.
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Publication Date: 2012-03-12 PubMed ID: 22411893DOI: 10.1002/cyto.a.22026Google Scholar: Lookup
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  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

The research article is about investigating cross-reactivity of commercial monoclonal antibodies on equine mesenchymal stem cells using flow cytometry. The established marker panel allows proper immunophenotypic characterization of horse mesenchymal stem cells.

Objective and Rationale

  • The objective of this study was to overcome the inherent challenges in the immunophenotypic characterization of equine MSCs due to the absence of a particular marker for these cells and limited availability of monoclonal antibodies (mAbs) specifically designed for them.
  • The researchers used a total of 30 commercial mAbs and flow cytometry to screen for those that can recognize equine epitopes, which are the parts of an antigen that are recognized by the immune system.

Process and Procedure

  • Multiple antibodies were assessed for their capacity to cross-react and identify horse mesenchymal stem cells. These included CD45, CD73, CD79α, CD90, CD105, MHC-II, a monocyte marker, CD29 and CD44.
  • Specificity of these antibodies was confirmed via confocal microscopy.
  • Four passages of undifferentiated equine MSCs derived from umbilical cord blood from six horses, were then characterized using flow cytometry and the nine-marker panel identified.
  • In order to selectively analyze viable, single cells, dead cells were excluded from analysis by the usage of a staining agent called 7-aminoactinomycin D.

Observations and Findings

  • The isolated equine MSCs from the fourth passage were discovered to be positive for CD29, CD44, CD90 and negative for CD45, CD79α, MHC-II, and a monocyte marker.
  • There was variable expression observed for CD73 and CD105.
  • The successful differentiation of these MSCs towards osteogenic, chondrogenic, and adipogenic lineage served as additional validation of their identity and characterization.

Conclusion

  • The study successfully established a nine-marker panel for the effective immunophenotyping of equine mesenchymal stem cells. This represents a significant forward step in identifying and isolating these cells for therapeutic applications in veterinary medicine.

Cite This Article

APA
De Schauwer C, Piepers S, Van de Walle GR, Demeyere K, Hoogewijs MK, Govaere JL, Braeckmans K, Van Soom A, Meyer E. (2012). In search for cross-reactivity to immunophenotype equine mesenchymal stromal cells by multicolor flow cytometry. Cytometry A, 81(4), 312-323. https://doi.org/10.1002/cyto.a.22026

Publication

Cytometry. Part A : the journal of the International Society for Analytical Cytology
ISSN: 1552-4930
NlmUniqueID: 101235694
Country: United States
Language: English
Volume: 81
Issue: 4
Pages: 312-323

Researcher Affiliations

De Schauwer, Catharina
  • Department of Reproduction, Obstetrics and Herd Health, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, Merelbeke 9820, Belgium. catharina.deschauwer@ugent.be
Piepers, Sofie
    Van de Walle, Gerlinde R
      Demeyere, Kristel
        Hoogewijs, Maarten K
          Govaere, Jan L J
            Braeckmans, Kevin
              Van Soom, Ann
                Meyer, Evelyne

                  MeSH Terms

                  • Adult
                  • Animals
                  • Antibodies, Monoclonal / immunology
                  • Cell Differentiation
                  • Cross Reactions
                  • Flow Cytometry / methods
                  • Horses / immunology
                  • Humans
                  • Immunophenotyping / methods
                  • Major Histocompatibility Complex / immunology
                  • Male
                  • Mesenchymal Stem Cells / cytology
                  • Mesenchymal Stem Cells / immunology
                  • Phenotype

                  Citations

                  This article has been cited 39 times.
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