Development and evaluation of a colorimetric membrane-array method for the detection of circulating tumor cells in the peripheral blood of Taiwanese patients with colorectal cancer.
Early detection is the hallmark of successful cancer treatment. Evidence is accumulating that primary cancers begin shedding neoplastic cells in the circulation at an early stage. To date, a high-sensitivity and high-throughput method for the detection of circulating tumor cells (CTCs) is deficient. In this study, we have developed a high-sensitivity colorimetric membrane-array method to detect CTCs in the peripheral blood of colorectal cancer (CRC) patients as a potential diagnostic tool. Previously, we identified a set of 18 oligonucleotide clones, significantly overexpressed in CRC, which were synthesized and applied to a nylon membrane. Digoxigenin (DIG)-labeled cDNA were amplified by reverse transcriptase-polymerase chain reaction (RT-PCR) from the peripheral blood of 88 Taiwanese CRC patients and 50 healthy subjects, and were then hybridized to the membrane-array. Hybridization signals were detected by color development. Meanwhile, blood samples were analyzed by real-time quantitative PCR (Q-PCR). Subsequently, both methods were compared regarding their correlation, sensitivity and specificity in the detection of CTCs by statistics. The results of membrane-arrays were demonstrated to be closely related to that of Q-PCR (P<0.001). The sensitivity and specificity of membrane-arrays for the detection of CTCs were 94.3% (95% CI, 86.4-102.2%) and 94% (95% CI, 85.9-102.1%), respectively. Moreover, the accuracy of membrane-arrays is higher than that of any one gene by Q-PCR. The detection rate of membrane-arrays was significantly associated with the depth of tumor invasion (P=0.002), the presence of lymph node metastasis (P=0.016), and TNM stage (P=0.005). The preliminary results indicated that the accuracy of membrane-arrays was sufficient to distinguish Taiwanese CRC patients from normal individuals with the advantages of time-saving, cost-effectiveness and high-throughput. Thus, the constructed colorimetric membrane-array could be a promising approach for the future detection of CTCs.
Wang
Jaw-Yuan
JY
Department of Surgery, Faculty of Medicine, College of Medicine, Kaohsiung Medical University Chung-Ho Memorial Hospital, Kaohsiung, Taiwan.
Yeh
Ching-Sheng
CS
Chen
Yi-Fang
YF
Wu
Chan-Hang
CH
Hsieh
Jan-Sing
JS
Huang
Tsung-Jan
TJ
Huang
Sung-Yu
SY
Lin
Shiu-Ru
SR
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D015179:Colorectal Neoplasms / Q000097:blood / Q000175:diagnosis* / Q000235:genetics; D004076:Digoxigenin / Q000737:chemistry; D005260:Female; D020869:Gene Expression Profiling / Q000295:instrumentation / Q000379:methods; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D006801:Humans; D016014:Linear Models; D008297:Male; D008875:Middle Aged; D009360:Neoplastic Cells, Circulating / Q000378:metabolism / Q000473:pathology*; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013624:Taiwan
D004076:Digoxigenin