Document Detail


Determination of cell concentration in a plant cell suspension using a fluorescence microplate reader.
MedLine Citation:
PMID:  15747158     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Microscopic counting of plant cells is a very tedious and time-consuming process and is therefore seldom used to evaluate plant cell number on a routine basis. This study describes a fast and simple method to evaluate cell concentration in a plant cell suspension using a fluorescence microplate reader. Eschscholtzia californica cells were fixed in a mix of methanol and acetic acid (3:1) and stained with a fluorescent DNA binding dye (Hoechst 33258). Readings were done in a fluorescence microplate reader at 360/465 nm. Specific binding of the dye to double-stranded DNA was significantly favored over unspecific binding when 1.0 M Tris buffer at pH 7.5 containing 1.0 M NaCl and 75 microg ml(-1) of Hoechst 33258 was used. Fluorescence readings must be done between 4 min and 12 min following the addition of the staining solution to the sample. The microplate counting method provides a convenient, rapid and sensitive procedure for determining the cell concentration in plant cell suspensions. The assay has a linear detection range from 0.2 x 10(6) cells to 10.0 x 10(6) cells per milliliter (actual concentration in the tested cell suspension). The time needed to perform the microplate counting was 10% of that needed for the microscopic enumeration. However, this microplate counting method can only be used on genetically stable cell lines and on asynchronous cell suspensions.
Authors:
Laurence Lamboursain; Mario Jolicoeur
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Publication Detail:
Type:  Evaluation Studies; Journal Article; Research Support, Non-U.S. Gov't     Date:  2004-12-10
Journal Detail:
Title:  Plant cell reports     Volume:  23     ISSN:  0721-7714     ISO Abbreviation:  Plant Cell Rep.     Publication Date:  2005 Mar 
Date Detail:
Created Date:  2005-03-04     Completed Date:  2005-06-16     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  9880970     Medline TA:  Plant Cell Rep     Country:  Germany    
Other Details:
Languages:  eng     Pagination:  665-72     Citation Subset:  IM    
Affiliation:
Canada Research Chair on the Development of Metabolic Engineering Tools, Bio-P2 Research Unit, Department of Chemical Engineering, Ecole Polytechnique de Montréal, P.O. Box 6079, Station Centre-ville, Montreal, Quebec, Canada, H3C 3A7.
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MeSH Terms
Descriptor/Qualifier:
Bisbenzimidazole* / metabolism
Cell Count*
Cells, Cultured
DNA, Plant / metabolism
Eschscholzia / cytology*,  growth & development
Fluorescent Dyes* / metabolism
Reproducibility of Results
Sensitivity and Specificity
Staining and Labeling / methods*
Chemical
Reg. No./Substance:
0/DNA, Plant; 0/Fluorescent Dyes; 23491-44-3/Bisbenzimidazole

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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