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

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.