Binary Assay For Bladder Cancer Analysis And Its Clinical Applications

Bladder cancer is the most common malignancies in urogenital system. Bladder cancer diagnosis and monitoring has been a big clinical problem. As we all know, cancer diagnosis is an indispensable precondition for the success of cancer disease treatment.Recently, the advantages of isothermal amplification have been mentioned repeatedly. Isothermal amplification has been used in the preparation of biosensors and the detection of biomarkers.After the PCR technology is realized,isothermal amplification, a new kind of technology for nucleic acid amplification in vitro technology, is flourished. Scientists from virous region are committed to apply the isothermal amplification technique to bladder cancer biomarkers detection in vitro. A large number of new tumor biomarkers have been discovered in recent years. However, the limitation of specific functions and detection methods of these biomarkers still keep this problem unresolved. Here we introduce a new assay with sensitivity, selectivity, and simplicity to meet the requirement of bladder cancer diagnosis and monitoring in clinical level. This assay not only do the optimization of traditional protein and nucleic acid detection, but also meet the clinical stability. The main contents of this article include:1. Recounted the application of isothermal amplification in the field of protein detection. Here we firstly demonstrate and verify the molecular beacon(MB) inhibiting the G-quadruplex to assist telomerase amplification. And, the key idea of the system is converting the biological signal to photochemical signals. We use GO and Nt.CviPII in this system to realize the amplification of detection signal. With detection limit of 0.4 cells/?L, the detection sensitivity of the system can reach the single-cell level successfully. Performance testes on about 60 clinical patients samples demonstrate that the detection rates of bladder cancer is around 70%, and false positive rate is reduced to 5.3%.2. Optimized the traditional RCA method to detect the miRNA, achieved the dual optimization of sensitivity and specificity. With the implementation of converting the biological signal to photochemical signals, we have a breakthrough in the field of clinical application. We also use GO and Nt.CviPII in the system to realize the amplification of detection signal. The detection limit of the system achieve 1pM. At the same time, the stability of the system is ensured. Performance testes on about 80 clinical patient samples demonstrate that the detection rates of bladder cancer is around 60% without false positive signals.3. We firstly demonstrate a binary assay to detect two different biomarkers of bladder cancer at the same time. And the system still possesses sensitivity, selectivity, and simplicity. And, with the achievement of simultaneously detection of miRNA and telomerase, binary assay can distinguish biomarkers from protein analogues, single-base and three base mismatch targets, which provides a new thought for miRNA sequence homology detection. Performance testes on about 70 urine samples from bladder cancer patients and normal individuals demonstrate that the sensitivity of binary assay is 96.9%, and the specificity is increased greatly(81.8%). The detection rates of bladder cancer is around 88.6%, and false positive rate is reduced to 5.3%. Crucially, this binary assay shows great potential to distinguish between muscle invasive bladder cancers(MIBCs) and non-muscle-invasive bladder cancers(NMIBC).