A novel staining protocol for multiparameter assessment of cell heterogeneity in Phormidium populations (cyanobacteria) employing fluorescent dyes
Jazyk angličtina Země Spojené státy americké Médium print-electronic
Typ dokumentu časopisecké články, práce podpořená grantem
PubMed
23437052
PubMed Central
PMC3577823
DOI
10.1371/journal.pone.0055283
PII: PONE-D-12-22206
Knihovny.cz E-zdroje
- MeSH
- barvení a značení metody MeSH
- časové faktory MeSH
- fluorescenční barviva metabolismus MeSH
- fluorescenční spektrometrie MeSH
- indoly metabolismus MeSH
- organické látky metabolismus MeSH
- sinice cytologie metabolismus MeSH
- tetrazoliové soli metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- 5-cyano-2,3-ditolyltetrazolium MeSH Prohlížeč
- DAPI MeSH Prohlížeč
- fluorescenční barviva MeSH
- indoly MeSH
- organické látky MeSH
- SYTOX Green MeSH Prohlížeč
- tetrazoliové soli MeSH
Bacterial populations display high heterogeneity in viability and physiological activity at the single-cell level, especially under stressful conditions. We demonstrate a novel staining protocol for multiparameter assessment of individual cells in physiologically heterogeneous populations of cyanobacteria. The protocol employs fluorescent probes, i.e., redox dye 5-cyano-2,3-ditolyl tetrazolium chloride, 'dead cell' nucleic acid stain SYTOX Green, and DNA-specific fluorochrome 4',6-diamidino-2-phenylindole, combined with microscopy image analysis. Our method allows simultaneous estimates of cellular respiration activity, membrane and nucleoid integrity, and allows the detection of photosynthetic pigments fluorescence along with morphological observations. The staining protocol has been adjusted for, both, laboratory and natural populations of the genus Phormidium (Oscillatoriales), and tested on 4 field-collected samples and 12 laboratory strains of cyanobacteria. Based on the mentioned cellular functions we suggest classification of cells in cyanobacterial populations into four categories: (i) active and intact; (ii) injured but active; (iii) metabolically inactive but intact; (iv) inactive and injured, or dead.
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