CAJ | 학술논문

目的建立定量检测小细胞肺癌(SCLC)循环肿瘤细胞(CTC)的方法,分析其敏感性和稳定性.方法设计前胃泌素释放肽原(preproGRP)特异性引物和探针,建立检测preproGRP mRNA的定量逆转录聚合酶链反应(RT-PCR)方法,通过癌细胞掺入法评价检测的敏感性.同时采用免疫磁珠法(MACS)分离纯化外周血CTC,结合形态学诊断计数CTC.结果 MACS分离外周血CTC的敏感性为30％左右,检出下限为5个CTC/ml全血.preproGRP定量RT-PCR检测外周血CTC的敏感性为0.64个CTC/反应,检出下限为50个CTC/ml全血,低于MACS,但通过Ct值计算的CTC数量与实际细胞数量符合程度(80％左右)高于MACS法.结论 preproGRP定量RT-PCR和MACS在检测SCLC患者外周血CTC方面各有优缺点,MACS敏感性更高,适合＜50个CTC/ml全血的检测,preproGRP定量RT-PCR检测的CTC数量更符合实际含量.
목적 건립정량검측소세포폐암(SCLC)순배종류세포(CTC)적방법,분석기민감성화은정성.방법 설계전위비소석방태원(preproGRP)특이성인물화탐침,건립검측preproGRP mRNA적정량역전록취합매련반응(RT-PCR)방법,통과암세포참입법평개검측적민감성.동시채용면역자주법(MACS)분리순화외주혈CTC,결합형태학진단계수CTC.결과 MACS분리외주혈CTC적민감성위30％좌우,검출하한위5개CTC/ml전혈.preproGRP정량RT-PCR검측외주혈CTC적민감성위0.64개CTC/반응,검출하한위50개CTC/ml전혈,저우MACS,단통과Ct치계산적CTC수량여실제세포수량부합정도(80％좌우)고우MACS법.결론 preproGRP정량RT-PCR화MACS재검측SCLC환자외주혈CTC방면각유우결점,MACS민감성경고,괄합＜50개CTC/ml전혈적검측,preproGRP정량RT-PCR검측적CTC수량경부합실제함량.
Objective To establish a quantitative method to detect circulating tumor cells (CTC) in patients with small cell lung cancer,and analyze its sensitivity and stability.Methods A specific primer and probe for prepro-gastrin-releasing peptide (preproGRP) was designed and a quantitative RT-PCR method was established to detect preproGRP mRNA.Cell incorporation method was used to evaluate the sensitivity.Magnetic cell sorting (MACS) was used to isolate and purify CTC from peripheral blood,and the MACS in combination with morphological diagnosis were used for cell counting.Results The isolation rate of CTC by MACS was 30％ and the lower detection limit was 5 cells per ml blood.The sensitivity of quantitative RT-PCR in detection of preproGRP mRNA in CTC was 0.64 cells per reaction,and the lower detection limit was 50 cells per ml blood,which was lower than that of MACS.However,the cell numbers calculated by Ct value was in greater accordance (about 80％) with actual cell numbers than that obtained by MACS.Conclusions PreproGRP quantitative RT-PCR and MACS have both advantages and disadvantages in detecting CTC of SCLC patients.MACS has a higher sensitivity,and is more favorable when CTC count is below 50 per ml blood.Meanwhile,preproGRP mRNA quantitative RT-PCR is more reliable in calculating actual cell numbers.