Description of a flow cytometry approach based on SYBR-14 staining for the assessment of DNA content, cell cycle analysis, and sorting of living normal and neoplastic cells.
BACKGROUND
This study aimed to expand the utilization of a simplified flow cytometric approach that employing SYBR-14/PI staining into broader flow cytometry applications, including (i) measurement of the DNA content; (ii) performing cell cycle analysis on mammalian cells; and (iii) sorting of live SYBR-14-stained mammalian cells based on DNA content.


METHODS
Cell lines of human origin were stained with SYBR-14 and propidium iodide (PI) and assessed by a dual-color flow cytometry. Finally, sorting of living SYBR-14-stained human cell lines was performed.


RESULTS
Dual staining with SYBR-14 and PI of human cells followed by flow cytometry analysis demonstrates that in addition to quality assessment, this staining could be utilized to determinate the DNA content on mammal cells. In addition, it resolves the diploid, tetraploid, and aneuploid DNA content. Furthermore, the SYBR-14-stained mammal cells were efficiently sorted based on DNA content and live cells were obtained. All these features have not been previously described with the utilization of this staining approach.


CONCLUSIONS
Results of this study demonstrate that this flow cytometric approach not only allows assessment of the viability of cells, but also the DNA content of mammal cells. In addition, this approach allows one to sort viable cells stained with SYBR-14. These findings open-up unexpected and unrestricted avenues for sorting of living mammal cells and provide significant advantages over the traditionally cumbersome sorting approaches for living cells, which demand very specialized and expensive UV light sources as well as sophisticated sorting procedures.
Nunez
R
R
Cancer Center Flow Cytometry Core Facility, University of Illinois at Chicago, Chicago, IL 60607, USA. rafaelnr@uic.edu
Garay
N
N
Villafane
C
C
Bruno
A
A
Lindgren
V
V
D002453:Cell Cycle; D045744:Cell Line, Tumor; D002469:Cell Separation; D002470:Cell Survival; D004247:DNA / Q000032:analysis*; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes; D006801:Humans; D007621:Karyotyping; D009369:Neoplasms / Q000473:pathology*; D009930:Organic Chemicals; D011003:Ploidies; D013194:Staining and Labeling
D005456:Fluorescent Dyes; D009930:Organic Chemicals; C093207:SYBR-14; D004247:DNA