title: 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.
abstract: 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.
authors.affiliation: Department of Surgery, Faculty of Medicine, College of Medicine, Kaohsiung Medical University Chung-Ho Memorial Hospital, Kaohsiung, Taiwan.
authors.forename: Jaw-Yuan
authors.initials: JY
authors.lastname: Wang
authors.forename: Ching-Sheng
authors.initials: CS
authors.lastname: Yeh
authors.forename: Yi-Fang
authors.initials: YF
authors.lastname: Chen
authors.forename: Chan-Hang
authors.initials: CH
authors.lastname: Wu
authors.forename: Jan-Sing
authors.initials: JS
authors.lastname: Hsieh
authors.forename: Tsung-Jan
authors.initials: TJ
authors.lastname: Huang
authors.forename: Sung-Yu
authors.initials: SY
authors.lastname: Huang
authors.forename: Shiu-Ru
authors.initials: SR
authors.lastname: Lin
mesh_terms: 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
chemical_list: D004076:Digoxigenin