Abstract
BackgroundFormalin has been widely used by pathology laboratories. Its carcinogenicity has led researchers to explore formalin substitutes. Streck Cell Preservative (SCP) is a formalin-free preservative that can preserve cellular antigens. This study was undertaken to investigate the effects of cell preservation using SCP on nucleic acid amplification, hybridization, and next-generation sequencing (NGS) as compared to control frozen cells and cells fixed in the traditional cell and tissue fixative, 10 % neutral buffered formalin (NBF).FindingsThe breast cancer cell line, SKBR-3, was used as a model system. Prior to nucleic acid extraction and fluorescence in situ hybridization (FISH), cells were fixed in SCP or NBF overnight at room temperature with frozen cells in parallel. Analysis showed that similar DNA extraction yields and amplification profiles determined by PCR in SCP preserved cells and control frozen cells, whereas NBF preserved cells had decreased DNA yield and impaired PCR amplification. Molecular cytogenetic studies by FISH technique indicated that the ratios of ERBB2 (HER-2/neu) signals to the chromosome 17 centromere (CEP17) were comparable for frozen cells and SCP preserved cells. The fluorescence images of both SCP fixed and control frozen cells were also clear and comparable. On the contrary, the same analysis was unsuccessful with NBF preserved cells due to poor hybridization quality. Our data also demonstrated that SCP had negligible effect on NGS testing.ConclusionWe conclude that SCP can be used as an alternative to NBF as a preservative for maintaining the integrity of nucleic acids for nucleic acid amplification, sequencing and FISH analysis.Electronic supplementary materialThe online version of this article (doi:10.1186/s13104-015-1725-4) contains supplementary material, which is available to authorized users.
Highlights
Formalin has been widely used by pathology laboratories
We conclude that Streck Cell Preservative (SCP) can be used as an alternative to neutral buffered formalin (NBF) as a preservative for maintaining the integrity of nucleic acids for nucleic acid amplification, sequencing and fluorescence in situ hybridization (FISH) analysis
Effect of fixatives on DNA yield and PCR amplification Cultured SKBR-3 cells were re-suspended in SCP (1:1 diluted with phosphate buffer solution (PBS)) or 10 % NBF and stored overnight at room temperature with unfixed frozen cells in parallel
Summary
Formalin has been widely used by pathology laboratories. Its carcinogenicity has led researchers to explore formalin substitutes. Streck Cell Preservative (SCP) is a formalin-free preservative that can preserve cellular antigens. This study was undertaken to investigate the effects of cell preservation using SCP on nucleic acid amplification, hybridization, and next-generation sequencing (NGS) as compared to control frozen cells and cells fixed in the traditional cell and tissue fixative, 10 % neutral buffered formalin (NBF). Preservation of cell and tissue morphology and antigenicity in clinical specimens is the foundation step in accurate diagnoses. Formalin preserved cells and tissues exhibit excellent morphology and acceptable antigenicity, but there are some major drawbacks. Studies show that formalin exerts a carcinogenic effect and poses health risks to the laboratory users [2].
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