Time-saving method for directly amplifying and capturing a minimal amount of pancreatic tumor-derived mutations from fine-needle aspirates using digital PCR
It is challenging to secure a cytopathologic diagnosis using minute amounts of tumor fluids and tissue fragments. Hence, we developed a rapid, accurate, low-cost method for detecting tumor cell-derived DNA from limited amounts of specimens and samples with a low tumor cellularity, to detect KRAS mut...
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Veröffentlicht in: | Scientific reports 2020-07, Vol.10 (1), p.12332-12332, Article 12332 |
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Sprache: | eng |
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Zusammenfassung: | It is challenging to secure a cytopathologic diagnosis using minute amounts of tumor fluids and tissue fragments. Hence, we developed a rapid, accurate, low-cost method for detecting tumor cell-derived DNA from limited amounts of specimens and samples with a low tumor cellularity, to detect
KRAS
mutations in pancreatic ductal carcinomas (PDA) using digital PCR (dPCR). The core invention is based on the suspension of tumor samples in pure water, which causes an osmotic burst; the crude suspension could be directly subjected to emulsion PCR in the platform. We examined the feasibility of this process using needle aspirates from surgically resected pancreatic tumor specimens (n = 12). We successfully amplified and detected mutant
KRAS
in 11 of 12 tumor samples harboring the mutation; the positive mutation frequency was as low as 0.8%. We used residual specimens from fine-needle aspiration/biopsy and needle flush processes (n = 10) for method validation. In 9 of 10 oncogenic
KRAS
pancreatic tumor samples, the "water-burst" method resulted in a positive mutation call. We describe a dPCR-based, super-sensitive screening protocol for determining
KRAS
mutation availability using tiny needle aspirates from PDAs processed using simple steps. This method might enable pathologists to secure a more accurate, minimally invasive diagnosis using minute tissue fragments. |
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ISSN: | 2045-2322 2045-2322 |
DOI: | 10.1038/s41598-020-69221-6 |