Ex vivo detection of apoptotic trophoblast cells applying flow cytofluorometry and immunocytochemistry using M30 antibody directed to the cytokeratin 18 neo-epitope

  • Authors:
    • Janna Krol
    • Karin Mengele
    • Irina Öttl-Mantchenko
    • Anita Welk
    • Irina Wasilewitsch
    • Stephanie Pildner von Steinburg
    • Karl-Theodor M. Schneider
    • Manfred Schmitt
  • View Affiliations

  • Published online on: September 1, 2005     https://doi.org/10.3892/ijmm.16.3.415
  • Pages: 415-420
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Abstract

Apoptosis of placental trophoblast cells has become the subject of intensive research. Recently, a monoclonal antibody (M30) directed against a neo-epitope of cytokeratin 18, that is formed after cleavage of this cytoskeletal protein by caspases, was shown to be of advantage over other tests for the detection of trophoblast cell apoptosis. In the present study, we describe a method for the enrichment of highly pure villous trophoblast cells based on the proteolytic digestion of placental tissue, density gradient separation of dissected cells, and immunoelimination of contaminating, non-trophoblast cells employing an antibody to the HLA class I antigen. The high purity (94-99%) of the trophoblast cell preparation was shown by antibody staining for cytokeratin 7 and absence of vimentin. Furthermore, we demonstrate that after a simple permeabilization and fixation step with 90% methanol and using the M30 CytoDeath, FITC-conjugated antibody, apoptotic trophoblast cells could be distinguished from non-apoptotic cells by flow cytofluorometry in a highly quantitative and sensitive fashion. Our protocol is an improvement over previously used methods such as immunocytochemistry as it allows to differentiate rapidly between competent and apoptotic trophoblast cells by the quantitative method of flow cytofluorometry.

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September 2005
Volume 16 Issue 3

Print ISSN: 1107-3756
Online ISSN:1791-244X

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Spandidos Publications style
Krol J, Mengele K, Öttl-Mantchenko I, Welk A, Wasilewitsch I, Pildner von Steinburg S, Schneider KM and Schmitt M: Ex vivo detection of apoptotic trophoblast cells applying flow cytofluorometry and immunocytochemistry using M30 antibody directed to the cytokeratin 18 neo-epitope. Int J Mol Med 16: 415-420, 2005
APA
Krol, J., Mengele, K., Öttl-Mantchenko, I., Welk, A., Wasilewitsch, I., Pildner von Steinburg, S. ... Schmitt, M. (2005). Ex vivo detection of apoptotic trophoblast cells applying flow cytofluorometry and immunocytochemistry using M30 antibody directed to the cytokeratin 18 neo-epitope. International Journal of Molecular Medicine, 16, 415-420. https://doi.org/10.3892/ijmm.16.3.415
MLA
Krol, J., Mengele, K., Öttl-Mantchenko, I., Welk, A., Wasilewitsch, I., Pildner von Steinburg, S., Schneider, K. M., Schmitt, M."Ex vivo detection of apoptotic trophoblast cells applying flow cytofluorometry and immunocytochemistry using M30 antibody directed to the cytokeratin 18 neo-epitope". International Journal of Molecular Medicine 16.3 (2005): 415-420.
Chicago
Krol, J., Mengele, K., Öttl-Mantchenko, I., Welk, A., Wasilewitsch, I., Pildner von Steinburg, S., Schneider, K. M., Schmitt, M."Ex vivo detection of apoptotic trophoblast cells applying flow cytofluorometry and immunocytochemistry using M30 antibody directed to the cytokeratin 18 neo-epitope". International Journal of Molecular Medicine 16, no. 3 (2005): 415-420. https://doi.org/10.3892/ijmm.16.3.415