| Cancer,also known as malignant tumor,is caused by the uncontrolled proliferation and metastasis of cells in the body after cell mutations.Cancer is still difficult to cure and has a very high mortality rate.Lung cancer is a general term for cancers originating in the trachea,bronchus and lung.It is reported that lung cancer is a major factor in cancer-related morbidity and mortality.In primary lung cancer,non-small cell lung cancer accounted for 85%,and small cell lung cancer accounted for 15%.According to statistics,the five-year relative survival rate of non-small cell lung cancer is 23%.The most fundamental reason for the low survival rate is that nearly two-thirds of lung cancers are diagnosed at an advanced stage.Therefore,early diagnosis can greatly improve the cure rate of lung cancer and prolong the survival time of patients.At present,the clinical diagnosis methods of lung cancer mainly include computed axial tomography,positron emission tomography,magnetic resonance imaging and transthoracic biopsy.However,these methods are expensive for most of the population,and require the use of large instruments and large hospitals,which cannot achieve a comprehensive screening of lung cancer.Therefore,it is urgent to develop a portable point-of-care(POCT)system that is available,highly sensitive,simple and easy to operate,and can achieve accurate diagnosis of lung cancer through simplified detection process.Liquid biopsy is a branch of in vitro diagnostic method.It can analyze relevant disease markers in a few body fluids in a non-invasive way to achieve preliminary diagnosis of disease and reduce the additional harm brought by traditional biopsy to patients.With a large number of studies on liquid biopsy methods,many simple and easy to operate methods have been gradually developed and used for the detection of biomarkers.Among them,the detection methods based on lateral flow strips and colorimetric have attracted extensive attention because the results are easy to distinguish.Among these,the lateral flow immunoassay strip analysis is one of the most widely used POCT tools.Due to its advantages of low cost,fast detection speed and good selectivity of target biomarkers,lateral flow strip analysis is widely used in portable biological detection.Based on the lung cancer markers in body fluids,such as exosomes and free miRNA,this paper designed and synthesized a series of simple and easy to operate detection systems using DNA,DSN enzyme,gold nanoparticles,etc.,for rapid and highly sensitive detection of biomarkers in body fluids.These detection systems include signal amplification,such as enzyme amplification,chain replication reaction amplification and hybridization substitution amplification,as well as signal output,such as test strip and colorimetric display.Specifically include:1.A lung cancer detection system based on exosomal miRNA detection with low cost,easy operation and high sensitivity was developed,which can be used for the immediate diagnosis of lung cancer.The detection system includes three parts: 1)Magnetic nanoparticles recognize,capture and enrich lung cancer specific exosomes;2)Signal amplification of exosomal miRNA-205 based on DSN enzyme;3)Read visual data with lateral flow test strip.The detection system realized the detection of exosomal miRNA in saliva and urine of lung cancer patients,and the detection limit was lower than 1 p M.The test results were verified to be consistent with the results of q RT-PCR of gold standard.The system can complete the detection of clinical saliva and urine samples in a short time,and realize the real non-invasive diagnosis of lung cancer.This research scheme will provide a new strategy for the preliminary and convenient diagnosis of lung cancer.2.A dual colorimetric lung cancer detection platform was developed to quickly and sensitively detect lung cancer related circulating tumor miRNA in serum samples.In this detection platform,the signal of miRNA in body fluid is quickly transformed into DNA signal by the hybrid chain reaction triggered by miRNA,which effectively reduces the interference caused by RNA instability.At the same time,the hybridization chain reaction of the system realizes the simultaneous signal amplification of two miRNAs,and quickly converts the signal into colorimetric signal.The platform detects two miRNAs at the same time,avoiding false positive results in cancer diagnosis.The colorimetric detection results that can be directly recognized by the naked eye make the application of the system easier.The dual colorimetric detection method successfully realized the detection of early lung cancer biomarkers miRNA-223 and miRNA-200 b in serum,and the detection limits were as low as 10 pm and 10 f M,respectively.This work provides a simple and feasible tool for accurate diagnosis of lung cancer.3.A dual miRNA detection platform based on triple-line lateral flow test strip was developed for the rapid and sensitive detection of multiple free lung cancer-related miRNAs in body fluid.In this system,the signals of the target miRNA are amplified and converted by the chain substitution reaction induced by miRNA to form the hybridization chain between biotin-modified DNA chain and TAMRA-modified DNA chain,and the hybridization chain between digoxin-modified DNA chain and FAMmodified DNA chain.One end of the two chains is combined with gold nanoparticles modified with anti-TAMRA antibody and anti-FAM antibody,respectively,and the other end is bound to the two detection bands of the triple-line strip.The gold aggregation makes the test paper colored to form a detection signal.This strategy realizes the qualitative detection of circulating tumor miRNA in lung cancer serum samples,and can distinguish normal people from cancer patients.4.A simultaneous detection platform of four miRNAs based on AND logic gate and chain replacement reaction was developed for precise and highly sensitive real-time detection of circulating tumor miRNA.The platform uses chain hybridization reaction to trigger the release of short strand DNA with signaling molecules.Two different short strand DNA and specific single strand DNA bind together through complementary pairing to generate signals on the test strip.Four miRNAs can trigger the signal output of two AND logic gates and generate two band signals on the test strip.The highly sensitive detection of four miRNAs mediated by this dual logic gate can greatly eliminate the false positive signal generated by the detection of a single target or a few targets,and achieve more accurate real-time diagnosis of lung cancer. |
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