MicroRNAs （miRNAs） are small non-coding RNAs that regulate gene expression at the post-transcriptional level, and their aberrant expression occurs during the development of malignant diseases. Recently, miRNAs have ...MicroRNAs （miRNAs） are small non-coding RNAs that regulate gene expression at the post-transcriptional level, and their aberrant expression occurs during the development of malignant diseases. Recently, miRNAs have been proposed as potential prognostic and predictive biomarkers for early diagnosis. However, a major obstacle in rapid miRNA analysis from real samples is the lack of ultrasensitive and quantitative techniques. In this regard, the use of chemiluminescence （CL） system offers a highly sensitive strategy for detecting miRNAs. In this article, an ultrasensitive approach has been established for the quantification ofmiRNAs, using magnetic beads （MBs） and alkaline phosphatase （AP）-based CL system. This technique depends on sandwich hybridization among MBs-labeled capture probes, target miRNAs and biotin-labeled reporter probes, conjugation of streptavidin-alkaline phosphatase （SA-AP） to biotin-labeled reporter probes, and CL detection of AP-linked targets. Detection of miR-21 with this technique demonstrated a high selectivity and an ultralow limit of detection （LOD） of 60 fM with an extraordinarily wide range of six orders of magnitudes. The quantitation could be achieved by direct detecting target miRNA in serum samples within a total time of 1.5 h and did not require reverse transcription and polymerase chain reaction （PCR） amplification. Therefore, this developed method shows great potential for early cancer diagnosis based on miRNAs as biomarkers.展开更多
基金supported by the National Key Program for Developing Basic Research (2014CB744501)the National High Technology Research and Development Program of China (2012AA022703)+8 种基金the National Key Special Science Program (2013ZX10004103-002)the National Natural Science Foundation of China (61471168, 61527806, 61271056)the Special Projects in Jiangsu Province (BL2014094)the Economical Forest Cultivation and Utilization of 2011 Collaborative Innovation Center in Hunan Province [(2013) 448]the Talents Planning of Six Summit Fields of Jiangsu Province (2013-WSN-056)Tianjin Medical University General Hospital Funding (ZYYFY2015029)China Postdoctoral Science Foundation funded project (2015T80487)Natural Science Foundation of Jiangsu Province (BK20140900)the Open Project of State Key Laboratory of Bioelectronics (2014HX12)
文摘MicroRNAs （miRNAs） are small non-coding RNAs that regulate gene expression at the post-transcriptional level, and their aberrant expression occurs during the development of malignant diseases. Recently, miRNAs have been proposed as potential prognostic and predictive biomarkers for early diagnosis. However, a major obstacle in rapid miRNA analysis from real samples is the lack of ultrasensitive and quantitative techniques. In this regard, the use of chemiluminescence （CL） system offers a highly sensitive strategy for detecting miRNAs. In this article, an ultrasensitive approach has been established for the quantification ofmiRNAs, using magnetic beads （MBs） and alkaline phosphatase （AP）-based CL system. This technique depends on sandwich hybridization among MBs-labeled capture probes, target miRNAs and biotin-labeled reporter probes, conjugation of streptavidin-alkaline phosphatase （SA-AP） to biotin-labeled reporter probes, and CL detection of AP-linked targets. Detection of miR-21 with this technique demonstrated a high selectivity and an ultralow limit of detection （LOD） of 60 fM with an extraordinarily wide range of six orders of magnitudes. The quantitation could be achieved by direct detecting target miRNA in serum samples within a total time of 1.5 h and did not require reverse transcription and polymerase chain reaction （PCR） amplification. Therefore, this developed method shows great potential for early cancer diagnosis based on miRNAs as biomarkers.
