The Optimization and Characterization of an RNA-Cleaving Fluorogenic DNAzyme Probe for MDA-MB-231 Cell Detection.
Breast cancer is one of the most frequently diagnosed cancers in females worldwide and lacks specific biomarkers for early detection. In a previous study, we obtained a selective RNA-cleaving Fluorogenic DNAzyme (RFD) probe against MDA-MB-231 cells, typical breast cancer cells, through the systematic evolution of ligands by exponential process (SELEX). To improve the performance of this probe for actual application, we carried out a series of optimization experiments on the pH value of a reaction buffer, the type and concentration of cofactor ions, and sequence minimization. The length of the active domain of the probe reduced to 25 nt from 40 nt after optimization, which was synthesized more easily and economically. The detection limit of the optimized assay system was 2000 MDA-MB-231 cells in 30 min, which is more sensitive than the previous one (almost 5000 cells). The DNAzyme probe was also capable of distinguishing MDA-MB-231 cell specifically from 3 normal cells and 10 other tumor cells. This probe with high sensitivity, selectivity, and economic efficiency enhances the feasibility for further clinical application in breast cancer diagnosis. Herein, we developed an optimization system to produce a general strategy to establish an easy-to-use DNAzyme-based assay for other targets.
Department of Chemistry, Tsinghua University, Beijing 100084, China. xpc14@mails.tsinghua.edu.cn.|The Ministry-Province Jointly Constructed Base for State Key Lab-Shenzhen Key Laboratory of Chemical Biology, the Graduate School at Shenzhen, Tsinghua University, Shenzhen, Guangdong 518055, China. xpc14@mails.tsinghua.edu.cn.
Pengcheng
P
Xue
The Ministry-Province Jointly Constructed Base for State Key Lab-Shenzhen Key Laboratory of Chemical Biology, the Graduate School at Shenzhen, Tsinghua University, Shenzhen, Guangdong 518055, China. eheshengnan@163.com.
Shengnan
S
He
Department of Chemistry, Tsinghua University, Beijing 100084, China. maoyu_gt@126.com.|The Ministry-Province Jointly Constructed Base for State Key Lab-Shenzhen Key Laboratory of Chemical Biology, the Graduate School at Shenzhen, Tsinghua University, Shenzhen, Guangdong 518055, China. maoyu_gt@126.com.
Yu
Y
Mao
The Ministry-Province Jointly Constructed Base for State Key Lab-Shenzhen Key Laboratory of Chemical Biology, the Graduate School at Shenzhen, Tsinghua University, Shenzhen, Guangdong 518055, China. dradow@163.com.
Long
L
Qu
Department of Chemistry, Tsinghua University, Beijing 100084, China. liu.feng@sz.tsinghua.edu.cn.|The Ministry-Province Jointly Constructed Base for State Key Lab-Shenzhen Key Laboratory of Chemical Biology, the Graduate School at Shenzhen, Tsinghua University, Shenzhen, Guangdong 518055, China. liu.feng@sz.tsinghua.edu.cn.
Feng
F
Liu
Department of Chemistry, Tsinghua University, Beijing 100084, China. tancy@sz.tsinghua.edu.cn.|The Ministry-Province Jointly Constructed Base for State Key Lab-Shenzhen Key Laboratory of Chemical Biology, the Graduate School at Shenzhen, Tsinghua University, Shenzhen, Guangdong 518055, China. tancy@sz.tsinghua.edu.cn.
Chunyan
C
Tan
Department of Chemistry, Tsinghua University, Beijing 100084, China. jiangyy@sz.tsinghua.edu.cn.|The Ministry-Province Jointly Constructed Base for State Key Lab-Shenzhen Key Laboratory of Chemical Biology, the Graduate School at Shenzhen, Tsinghua University, Shenzhen, Guangdong 518055, China. jiangyy@sz.tsinghua.edu.cn.
Yuyang
Y
Jiang
Department of Chemistry, Tsinghua University, Beijing 100084, China. tan.ying@sz.tsinghua.edu.cn.|The Ministry-Province Jointly Constructed Base for State Key Lab-Shenzhen Key Laboratory of Chemical Biology, the Graduate School at Shenzhen, Tsinghua University, Shenzhen, Guangdong 518055, China. tan.ying@sz.tsinghua.edu.cn.
Ying
Y
Tan
D001943:Breast Neoplasms; D021881:DNA, Catalytic / Q000378:metabolism*; D005260:Female; D006801:Humans; D007477:Ions; D057230:Limit of Detection; D015347:RNA Probes
D021881:DNA, Catalytic; D007477:Ions; D015347:RNA Probes
DNAzyme; MDA-MB-231 cell; RFD probe; optimization