Das Kausik, Fernando M Rohan, Basiaga Sara, Wigginton Stephanie M, Williams Tom
Streck, Inc., 7002 S 109 Street, La Vista, NE 68128, USA.
Department of Chemistry, University of Nebraska, Lincoln, NE 68588, USA.
Acta Histochem. 2014 Jan;116(1):55-60. doi: 10.1016/j.acthis.2013.05.002. Epub 2013 Jun 28.
Stabilization of nucleated blood cells by cell stabilizing reagent (BCT reagent) present in the Cell-Free DNA BCT blood collection device and consequent prevention of cell-free DNA contamination by cellular DNA during sample storage and shipping have previously been reported. This study was conducted to investigate the effect of this novel cell stabilizing reagent on DNA amplification by PCR as compared to traditional cell stabilizing reagents, formaldehyde and glutaraldehyde. A 787 bp long DNA fragment from human glyceraldehydes-3-phosphate dehydrogenase (GAPDH) gene was amplified by PCR and used as model system. DNA samples and blood samples were treated with BCT reagent, 0.1% formaldehyde or 0.1% glutaraldehyde at room temperature. DNA amplification was studied using conventional and real-time quantitative PCR. Results indicate that exposure of DNA to the BCT reagent for up to 14 days had no effect on DNA amplification by PCR as compared to the untreated control DNA. However, there was statistically significant decrease in DNA amplification in the DNA samples treated with formaldehyde and glutaraldehyde. We conclude that the BCT reagent used in Cell-Free DNA BCT blood collection device to prevent cell-free DNA contamination by cellular DNA had no effect on DNA amplification by PCR.
先前已有报道，无细胞DNA BCT采血装置中存在的细胞稳定试剂（BCT试剂）可使有核血细胞稳定，并在样本储存和运输过程中防止细胞DNA污染无细胞DNA。本研究旨在探讨这种新型细胞稳定试剂与传统细胞稳定试剂甲醛和戊二醛相比，对PCR扩增DNA的影响。通过PCR扩增来自人甘油醛-3-磷酸脱氢酶（GAPDH）基因的787 bp长的DNA片段，并将其用作模型系统。在室温下，用BCT试剂、0.1%甲醛或0.1%戊二醛处理DNA样本和血液样本。使用常规PCR和实时定量PCR研究DNA扩增。结果表明，与未处理的对照DNA相比，DNA暴露于BCT试剂长达14天对PCR扩增DNA没有影响。然而，用甲醛和戊二醛处理的DNA样本中，DNA扩增有统计学意义的下降。我们得出结论，无细胞DNA BCT采血装置中用于防止细胞DNA污染无细胞DNA的BCT试剂对PCR扩增DNA没有影响。
