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DNA extraction from archived hematoxylin and eosin-stained tissue slides for downstream molecular analysis

World Journal of Methodology 2019年第3期9卷 32-43页

作者： Pushkal sinduvadi ramesh Venkatesh Madegowda suprith Kumar shailashree Narasimha parichay s r Nandini Nandish Manoli Devananda Devegowda Center of Excellence in Molecular Biology and regenerative Medicine Department of BiochemistryJSS Medical CollegeJSS Academy of Higher Education and ResearchMysuru 570015India CIPHEr Healthcare Pvt Ltd. Hyderabad 500034India Department of Pathology JSS Medical CollegeJSS Academy of Higher Education and ResearchMysuru 570015India

BACKGrOUND Histopathologically stained archived tissue slides are stored in hospital archives for years to *** are the largest available source of biological materials and are a potentially useful resource that can be used for retrospective epidemiological *** recovered from the slides can be used for several downstream molecular processes including polymerase chain reaction,single nucleotide polymorphism analysis,and whole genome *** DNA from these slides can be utilized to compare gene signatures of normal and diseased ***,extraction of high-quality DNA from archived stained hematoxylin and eosin(H&E)slides remains *** To standardize a new protocol for extracting DNA from archived H&E-stained tissue slides for further molecular *** A total of 100 archived H&E-stained cancer slides were subjected to a total of five methods of DNA *** were varied in the deparaffinization step,tissue rehydration,duration of lysis,and presence or absence of proteinase *** extracted DNA was quantified using a NanoDrop spectrophometer and the quality was analyzed by agarose gel *** each sample was subjected to polymerase chain reaction(PCr)to amplify the internal control gene GAPDH,thereby confirming the DNA intactness,which could be further utilized for other downstream *** Of the five different methods tested,the third method wherein xylene was used for tissue deparaffinization followed by 72 h of digestion and without proteinase K inactivation yielded the highest amount of DNA with good *** yield was significantly higher when compared to other *** addition,90%of the extracted DNA showed amplifiable GAPDH *** Here we present a step-by-step,cost-effective,and reproducible protocol for the extraction of PCr-friendly DNA from archived H&E-stained cancer tissue slides that can be used for further downstream molecular applications.

关键词： DNA extraction Hematoxylin and eosin tissue slides Molecular analysis Polymerase chain reaction Deparaffinization

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