Circulating tumor cells (CTCs) from peripheral blood are emerging as a useful tool for the detection of malignancy, monitoring disease progression, and measuring response to therapy. We describe a unique microfluidic chip that was capable of efficient and selective separation of CTCs from peripheral whole blood samples. The ability of microfluidic chip to capture CTCs from PBS and whole blood samples was tested.Sixty-eight peripheral blood samples from 68 colorectal cancer patients were investigated for the presence of CTCs by microchip technology. The frequency of CTCs was analyzed statistically for correlation with relevant clinical data. We also examined samples from 20 healthy individuals as controls. The calculated capture efficiency was 85.7% and decreased significantly at flow rates above 2.0 ml/h. The number of CTCs isolated ranged from 3 to 236/ml for colorectal patients [99 ± 64 (mean ± SD) CTCs/ml]. None of the 20 healthy subjects had any identifiable CTCs. We identified CTCs in 46 (67.65%) of the 68 patients: in two of nine (22.22%) Dukes A, in 10 of 24 (41.67%) Dukes B, in 21 of 22 (95.45%) Dukes C, and in all 13 Dukes D patients. The detection rate in Dukes C and D patients was much higher than in Dukes A and B patients (97.73% vs. 36.36%) (p < 0.01). A significant correlation between detection of CTCs and clinical stage (r = 0.792, p < 0.01) was found, which was higher than carcinoembryonic antigen (r = 0.285, p > 0.01), carbohydrate antigen 19-9 (r = 0.258, p > 0.01), α-fetoprotein (r = 0.096, p > 0.01), and cancer antigen 125 (r = 0.134, p > 0.01). Microfluidic chip provides a novel method for capturing CTCs. The presence of CTCs correlated with clinical stage. It is important to evaluate CK-positive and DAPI-stained tumor cells together to determine the role of CTCs in tumor behavior and disease progression.