NIH, National Cancer Institute, Division of Cancer Treatment and Diagnosis (DCTD) NIH - National Institutes of Health National Cancer Institute DCTD - Division of Cancer Treatment and Diagnosis
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Extraction of DNA from exfoliative cytology specimens and its suitability for analysis by the polymerase chain reaction.

Author(s): Jackson DP, Payne J, Bell S, Lewis FA, Taylor GR, Peel KR, Sutton J, Quirke P

Publication: Cytopathology, 1990, Vol. 1, Page 87-96

PubMed ID: 1966323 PubMed Review Paper? No

Purpose of Paper

The purpose of this paper was to assess the utility of fresh and archival exfoliative cytology samples for use in the polymerase chain reaction (PCR).

Conclusion of Paper

The authors report that both fresh and archival cervical cytology specimens are suitable for DNA extraction and subsequent amplification with PCR. Proteinase K digestion for 1 hour resulted in the greatest yield of high quality DNA, which was successfully analyzed by PCR. Of note, the other extraction methods evaluated, which included SDS incubation and specimen boiling, also resulted in successful PCR analysis although DNA yields and fragment size were somewhat lower.

Studies

  1. Study Purpose

    The purpose of this study was to optimize methods of DNA extraction from cytology specimens for PCR amplification of the KM19, N-ras, and K-ras genes and the Alu multiple copy repeat sequence. Cervical cells as well as cells collected from sputa, urine, pleural effusions, and ascitic fluid were examined.

    Summary of Findings:

    DNA extraction with a 1 h proteinase K incubation resulted in the greatest yield for both fresh and archival cervical exfoliative cytology specimens compared to the other methods of protein digestion evaluated. DNA yields from archived specimens other than cervix were variable, but generally were low and comprised of low molecular weight DNA. PCR amplification was successful for fresh and archival cervical specimens for all extraction methods investigated. PCR analysis of archived specimens other than cervix produced variable and primer-specific results.

    Biospecimens
    Preservative Types
    • None (Fresh)
    Diagnoses:
    • Not specified
    Platform:
    AnalyteTechnology Platform
    DNA Electrophoresis
    DNA Spectrophotometry
    DNA PCR
    Pre-analytical Factors:
    ClassificationPre-analytical FactorValue(s)
    Analyte Extraction and Purification Protein digestion 0.1 mg/ml proteinase K, 1 h, 50 degrees C
    0.1 mg/ml proteinase K, 1 h, 37 degrees C
    0.1 mg/ml proteinase K, 18 h, 37 degrees C
    1% SDS, 1 h, 50 degrees C
    Boiling for 15 min
    Storage Storage duration 7 years
    PCR Specific Type of tissue stain Papanicolaou
    Biospecimen Acquisition Biospecimen location Cervix
    Sputa
    Urine
    Pleural effusion
    Ascitic fluid
    PCR Specific Targeted nucleic acid KM19
    N-ras
    K-ras
    Alu repeats
    View more

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