Nunez R, Garay N, Villafane C, Bruno A, Lindgren V
Cancer Center Flow Cytometry Core Facility, University of Illinois at Chicago, Chicago, IL 60607, USA.
Exp Mol Pathol. 2004 Feb;76(1):29-36. doi: 10.1016/j.yexmp.2003.10.004.
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.
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.
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.
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.
本研究旨在将采用SYBR-14/PI染色的简化流式细胞术方法扩展到更广泛的流式细胞术应用中，包括：（i）测量DNA含量；（ii）对哺乳动物细胞进行细胞周期分析；（iii）基于DNA含量对SYBR-14染色的活哺乳动物细胞进行分选。
用人源细胞系进行SYBR-14和碘化丙啶（PI）染色，并通过双色流式细胞术进行评估。最后，对SYBR-14染色的活人细胞系进行分选。
人细胞用SYBR-14和PI进行双色染色，随后进行流式细胞术分析，结果表明，除了质量评估外，这种染色还可用于测定哺乳动物细胞的DNA含量。此外，它还能分辨二倍体、四倍体和非整倍体的DNA含量。此外，基于DNA含量对SYBR-14染色的哺乳动物细胞进行了有效分选，并获得了活细胞。利用这种染色方法，所有这些特性以前都没有被描述过。
本研究结果表明，这种流式细胞术方法不仅可以评估细胞活力，还可以评估哺乳动物细胞的DNA含量。此外，这种方法还可以对SYBR-14染色的活细胞进行分选。这些发现为活哺乳动物细胞的分选开辟了意想不到且不受限制的途径，与传统上繁琐的活细胞分选方法相比具有显著优势，传统方法需要非常专业且昂贵的紫外光源以及复杂的分选程序。
