Dna Flow Cytometry of Breast Carcinoma After Acetic-Acid Fixation

Abstract

Aqueous acetic acid was used to fix and store specimens of [human] tissue prior to dissociation into nuclear suspensions for flow cytometric quantitation of DNA. The optimum concentration was 20 vol of glacial acetic acid in 80 vol of distilled water. Both neoplastic and benign nuclei were easily released from the fixed tissue blocks by slicing and shaking. Residual undissociated tissue was suitable for microscopic examination to confirm its identity. The nuclei fluoresced brightly after staining with propidium iodide, and yielded histograms similar to those from unfixed samples. Acetic-acid fixation resulted in slightly broader G1 and G0 peaks in the DNA histograms in comparison to unfixed cells, but fluorescent debris was less and correlation between the flow cytometric S-phase fraction (SPF) and in-vitro thymidine labeling index (TLI) was better than with unfixed cells. Twenty-one of 39 acetic-acid-fixed breast carcinomas had DNA indices > 1.0 (increased nuclear DNA content in comparison to benign cells), and 18 had DNA indices of 1.0 (normal or near-normal). The SPF was usually in excess of the TLI, but the 2 were significantly correlated (r = 0.72, P < 0.0001). A significant correlation of SPF with TLI held only for the group with DNA index > 1.0. DNA indices > 1.0 were associated with high SPF and TLI, and high SPF and TLI each associated with low content of estrogen receptor. [Therapeutic, prognostic indications are considered].