Zhenhuan Jin, Wei Xiao, Lin Shen, Xiaoxue Shi, Jianping Li
{"title":"一种基于 CRISPR-Cas12a 和酶反应的电化学方法,用于高灵敏度检测肿瘤标志物 MUC1 粘蛋白","authors":"Zhenhuan Jin, Wei Xiao, Lin Shen, Xiaoxue Shi, Jianping Li","doi":"10.1039/d4an00595c","DOIUrl":null,"url":null,"abstract":"The early warning of cancer is crucial in cancer prevention and anti-cancer, and the highly sensitive methods for detecting cancer biomarkers are essential for cancer early diagnostic. Herein, an electrochemical aptamer biosensor based on CRISPR-Cas12a system was constructed for the detection of cancer tumor biomarker MUC1 mucin. The sensitivity was significantly prompted by enzyme-catalyzed signal amplification, and the selectivity was improved by dual recognition of the aptamer to MUC1 and the crRNA-Cas12a system to aptamer. Glucose oxidase (GOD) was loaded on the surface of magnetic Fe3O4@Au (MGNP) via probe single-stranded DNA (pDNA) with terminal modification of mercapto (-SH) to form GOD-pDNA/MGNP. The corresponding aptamer of MUC1 (MUC1 Apt) binds to its complementary ssDNA (cDNA) to form the activator Apt/cDNA, which is specifically recognized by crRNA-Cas12a and excites the trans-cleavage function of Cas12a, thus in turn trans-cleave pDNA and detached GOD from the magnetic particles. The magnetic beads were separated and transferred into a glucose solution, and the oxidation current of H2O2 produced by the catalytic reaction of GOD was measured on a Pt modified magnetically controlled glassy carbon electrode, resulting in an indirect determination of MUC1. The current change was linear with the logarithm of MUC1 concentration in the range of 1.0×10-17 g/mL to 1.0×10-10 g/mL. The detection limit was as low as 7.01×10-18 g/mL. The method has been applied to the detection of MUC1 in medical samples.","PeriodicalId":63,"journal":{"name":"Analyst","volume":null,"pages":null},"PeriodicalIF":3.6000,"publicationDate":"2024-06-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"An electrochemical method based on CRISPR-Cas12a and enzymatic reaction for highly sensitive detection of tumor marker MUC1 mucin\",\"authors\":\"Zhenhuan Jin, Wei Xiao, Lin Shen, Xiaoxue Shi, Jianping Li\",\"doi\":\"10.1039/d4an00595c\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"The early warning of cancer is crucial in cancer prevention and anti-cancer, and the highly sensitive methods for detecting cancer biomarkers are essential for cancer early diagnostic. Herein, an electrochemical aptamer biosensor based on CRISPR-Cas12a system was constructed for the detection of cancer tumor biomarker MUC1 mucin. The sensitivity was significantly prompted by enzyme-catalyzed signal amplification, and the selectivity was improved by dual recognition of the aptamer to MUC1 and the crRNA-Cas12a system to aptamer. Glucose oxidase (GOD) was loaded on the surface of magnetic Fe3O4@Au (MGNP) via probe single-stranded DNA (pDNA) with terminal modification of mercapto (-SH) to form GOD-pDNA/MGNP. The corresponding aptamer of MUC1 (MUC1 Apt) binds to its complementary ssDNA (cDNA) to form the activator Apt/cDNA, which is specifically recognized by crRNA-Cas12a and excites the trans-cleavage function of Cas12a, thus in turn trans-cleave pDNA and detached GOD from the magnetic particles. The magnetic beads were separated and transferred into a glucose solution, and the oxidation current of H2O2 produced by the catalytic reaction of GOD was measured on a Pt modified magnetically controlled glassy carbon electrode, resulting in an indirect determination of MUC1. The current change was linear with the logarithm of MUC1 concentration in the range of 1.0×10-17 g/mL to 1.0×10-10 g/mL. The detection limit was as low as 7.01×10-18 g/mL. The method has been applied to the detection of MUC1 in medical samples.\",\"PeriodicalId\":63,\"journal\":{\"name\":\"Analyst\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":3.6000,\"publicationDate\":\"2024-06-11\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Analyst\",\"FirstCategoryId\":\"92\",\"ListUrlMain\":\"https://doi.org/10.1039/d4an00595c\",\"RegionNum\":3,\"RegionCategory\":\"化学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"CHEMISTRY, ANALYTICAL\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Analyst","FirstCategoryId":"92","ListUrlMain":"https://doi.org/10.1039/d4an00595c","RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"CHEMISTRY, ANALYTICAL","Score":null,"Total":0}
An electrochemical method based on CRISPR-Cas12a and enzymatic reaction for highly sensitive detection of tumor marker MUC1 mucin
The early warning of cancer is crucial in cancer prevention and anti-cancer, and the highly sensitive methods for detecting cancer biomarkers are essential for cancer early diagnostic. Herein, an electrochemical aptamer biosensor based on CRISPR-Cas12a system was constructed for the detection of cancer tumor biomarker MUC1 mucin. The sensitivity was significantly prompted by enzyme-catalyzed signal amplification, and the selectivity was improved by dual recognition of the aptamer to MUC1 and the crRNA-Cas12a system to aptamer. Glucose oxidase (GOD) was loaded on the surface of magnetic Fe3O4@Au (MGNP) via probe single-stranded DNA (pDNA) with terminal modification of mercapto (-SH) to form GOD-pDNA/MGNP. The corresponding aptamer of MUC1 (MUC1 Apt) binds to its complementary ssDNA (cDNA) to form the activator Apt/cDNA, which is specifically recognized by crRNA-Cas12a and excites the trans-cleavage function of Cas12a, thus in turn trans-cleave pDNA and detached GOD from the magnetic particles. The magnetic beads were separated and transferred into a glucose solution, and the oxidation current of H2O2 produced by the catalytic reaction of GOD was measured on a Pt modified magnetically controlled glassy carbon electrode, resulting in an indirect determination of MUC1. The current change was linear with the logarithm of MUC1 concentration in the range of 1.0×10-17 g/mL to 1.0×10-10 g/mL. The detection limit was as low as 7.01×10-18 g/mL. The method has been applied to the detection of MUC1 in medical samples.