New applications of the acridine orange fluorescence staining method: Screening for circulating tumor cells

Abstract

The aim of the present study was to explore use of the acridine orange fluorescence (AO-F) staining method for screening of circulating tumor cells (CTCs) in renal cell carcinoma (RCC) patients. The AO-F positive staining rate of live and dead tumor cells was calculated. The positive staining rate in the live group was 93.4±3.0%, while the dead group failed to emit specific fluorescence. A known number of tumor cells were added to peripheral blood, and the detection sensitivity of the four groups (50, 100, 200 and 500 cells/tube) was 10.2±3.8, 9.2±2.3, 10.8±2.6 and 10.5±1.9%, respectively. The average detection sensitivity of the four groups was 10.16±2.73%. There was a positive correlation between the number of cells that was positively stained with AO‑F and the total number cells in the system (χ2=0.959; P<0.001). Subsequently, the AO‑F staining method was used to detect positive staining cells in 8 healthy volunteers (control group), and 112 non‑metastatic and 27 metastatic RCC patients. The positive staining rate was 13.67% (19/139) in RCC patients, while none of the control group was positive. The AO‑F positive staining rate was not significantly different between the metastatic and non‑metastatic patients according to age, gender, the pathological pattern, T2/3 (according to the Tumor‑Node‑Metastasis classification) or Fuhrman grade, while there was a significant difference according to T1. The positive staining rate was 8.93% (10/112) for non‑metastatic patients and 33.33% (9/27) for metastatic patients, which showed a significant difference (P<0.05). In 112 non‑metastatic and 27 metastatic patients, the positive staining rate was not significantly associated with gender, age, tumor size, the pathological pattern, T classification, Fuhrman grade, the presence of a lesion or metastasis to the lungs. The present study demonstrated that the method of CTC staining with AO‑F, which has high reproducibility and specificity, was feasible for identifying CTCs and warrants further study.