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Phenotypic and functional characterization of earthworm coelomocyte subsets: Linking light scatter-based cell typing and imaging of the sorted populations

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  • Péter Engelmann, University of Pecs, Hungary
  • Yuya Hayashi
  • Kornélia Bodo, University of Pecs, Hungary
  • David Ernszt, University of Pecs, Janos Szentagothai Research Centre, Hungary
  • Ildiko Somogyi, University of Pecs, Hungary
  • Anita Steib, University of Pecs, Hungary
  • Jozsef Orban, Janos Szentagothai Research Centre, Hungary
  • Edit Pollak, University of Pecs, Hungary
  • Miklos Nyitrai, University of Pecs, Janos Szentagothai Research Centre, MTA-PTE Nuclear-Mitochondrial Interactions Research Group, Hungary
  • Péter Németh, University of Pecs, Hungary
  • Laszlo Molnar, University of Pecs, Hungary
Flow cytometry is a common approach to study invertebrate immune cells including earthworm coelomocytes. However, the link between light-scatter- and microscopy-based phenotyping remains obscured. Here we show, by means of light scatter-based cell sorting, both subpopulations (amoebocytes and eleocytes) can be physically isolated with good sort efficiency and purity confirmed by downstream morphological and cytochemical applications. Immunocytochemical analysis using anti-EFCC monoclonal antibodies combined with phalloidin staining has revealed antigenically distinct, sorted subsets. Screening of lectin binding capacity indicated wheat germ agglutinin (WGA) as the strongest reactor to amoebocytes. This is further evidenced by WGA inhibition assays that suggest high abundance of N-acetyl-d-glucosamine in amoebocytes. Post-sort phagocytosis assays confirmed the functional differences between amoebocytes and eleocytes, with the former being in favor of bacterial engulfment. This study has proved successful in linking flow cytometry and microscopy analysis and provides further experimental evidence of phenotypic and functional heterogeneity in earthworm coelomocyte subsets.
Original languageEnglish
JournalDevelopmental & Comparative Immunology
Pages (from-to)41-52
Number of pages12
Publication statusPublished - 24 Jun 2016

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