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Cell Cycle Analysis; DNA Ploidy and S phase

PLOIDY

Test Synonym:

None

Test Synonym:

88182 – Flow cytometry, DNA or cell cycle analysis

Turnaround Time:

3 days

Methodology:

Flow cytometry analysis

Specimen Requirements:

  • 5.0ml (min. 2.0 ml) heparin, EDTA or ACD whole blood
  • 3.0mL (min. 1.0mL) bone marrow
  • 5mm3 fresh or frozen tissue in MPLN RPMI
  • Formalin-fixed paraffin embedded tissue
  • 30.0ml (min. 15.0ml) fresh urine in sterile container

    • Causes for Rejection:

    Insufficient volume; fresh samples not sent within 24 hours


    Specimen Stability:

  • Whole blood or bone marrow stable up to 48 hours 18- 25°C
  • Fresh tissue stable at 2-8°C up to 72 hours
  • Frozen tissue stable indefinitely at -20°C
  • Formalin-fixed paraffin embedded tissue stable at room temperature indefinitely
  • Fresh urine stable up to 48 hours at 2-8°C

    • Storage and Handling:

  • Whole blood, bone marrow and formalin fixed paraffin embedded tissue ship ambient
  • Fresh tissue and urine ship in a styrofoam container with an ice pack (do not allow the ice pack to directly contact the sample)
  • Frozen specimens should be shipped on dry ice

    • Reference Range:

    Diploid population detected

    Indication:

    Deoxyribonucleic acid (DNA) ploidy and cell cycle analysis is a rapid and efficient way to evaluate the DNA content (ploidy) and proliferative activity (cell cycle) of malignant cells. By staining the DNA with a fluorescent dye, flow cytometry can accurately measure the dye fluorescence in many individual cells, resulting in a histogram of DNA contents within the entire population. These data can be analyzed for proliferative activity (S-phase fraction) and diploid (normal content) or aneuploid (abnormal content). Ploidy has been shown to be a prognostic indicator for malignancy in many solid tumors based on the fact that malignant cells sometimes show abnormal chromosome numbers and the frequency of these abnormalities often increases as the tumor progresses to a higher grade. The S-phase fraction (SPF) or % S-phase refers to the percent or proportion of cells preparing for mitosis by their active synthesis of DNA. Tumor cells tend to replicate faster than normal cells increasing their SPF. In general, a high SPF correlates positively with poor differentiation, increasing tumor size and degrees of tumor spread, which all have prognostic significance.

    A quantitative assessment of DNA ploidy in tumors is achieved by noting the DNA index (DI), which is the ratio of the mean tumor sample G0/G1 DNA content compared to that of normal diploid reference cells. A normal or malignant diploid cell would have a DI of 1. The greater the deviation of the DI from 1 the more “aneuploid” the tumor.

    In conclusion, several studies have shown correlation between DNA ploidy, S-phase proliferation and tumor progression. Tumors from prostate, melanoma, breast carcinoma and bladder have been analyzed for ploidy and SPF. Tumor size and DNA synthetic activity were found to be directly related to clinical outcome in these malignancies.

    References:

    1. Tinari et al. (1993). DNA and S-phase fraction analysis by flow cytometry in prostate cancer. Clinicopathologic implications. Cancer  71(4):1289
    2. Martin et al. (2000). Value of DNA ploidy and S-phase fraction as prognostic factors in stage III cutaneous melanoma. Can J Surg 43(1):29
    3. Pinto et al. (1999). Short-term significance of DNA ploidy and cell proliferation in breast carcinoma: a multivariate analysis of prognostic markers in a series of 308 patients. J Clin Pathol 52(8):604