Document Detail

Viable cell sorting of dinoflagellates by multiparametric flow cytometry.
MedLine Citation:
PMID:  20305733     Owner:  NLM     Status:  Publisher    
Electronic cell sorting for isolation and culture of dinoflagellates and other marine eukaryotic phytoplankton was compared to the traditional method of manually picking cells using a micropipette. Trauma to electronically sorted cells was not a limiting factor, as fragile dinoflagellates, such as Karenia brevis (Dinophyceae), survived electronic cell sorting to yield viable cells. The rate of successful isolation of large-scale (> 4 litres) cultures was higher for manual picking than for electronic cell sorting (2% vs 0.5%, respectively). However, manual picking of cells is more labor intensive and time consuming. Most manually isolated cells required repicking, as the cultures were determined not to be unialgal after a single round of isolation; whereas, no cultures obtained in this study from electronic single-cell sorting required resorting. A broad flow cytometric gating logic was employed to enhance species diversity. The percentages of unique genotypes produced by manual picking or electronic cell sorting were similar (57% vs 54%, respectively), and each approach produced a variety of dinoflagellate or raphidophyte genera. Alternatively, a highly restrictive gating logic was successfully used to target K. brevis from a natural bloom sample. Direct electronic single-cell sorting was more successful than utilizing a pre-enrichment sort followed by electronic single-cell sorting. The appropriate recovery medium may enhance the rate of successful isolations. Seventy percent of isolated cells were recovered in a new medium (RE) reported here, which was optimized for axenic dinoflagellate cultures. The greatest limiting factor to the throughput of electronic cell sorting is the need for manual postsort culture maintenance and assessment of the large number of isolated cells. However, when combined with newly developed automated methods for growth screening, electronic single-cell sorting has the potential to accelerate the discovery of new algal strains.
Christopher D Sinigalliano; Jamie Winshell; Maria A Guerrero; Gloria Scorzetti; Jack W Fell; Richard W Eaton; Larry Brand; Kathleen S Rein
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Publication Detail:
Journal Detail:
Title:  Phycologia     Volume:  48     ISSN:  0031-8884     ISO Abbreviation:  -     Publication Date:  2009 Jul 
Date Detail:
Created Date:  2010-7-13     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0046550     Medline TA:  Phycologia     Country:  -    
Other Details:
Languages:  ENG     Pagination:  249-257     Citation Subset:  -    
Southeast Environmental Research Center, Florida International University, Miami, FL 33199, USA.
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Grant Support
P50 ES012736-05//NIEHS NIH HHS

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