RESUMEN
Anti-cancer therapy is crucial in cancer prevention and anti-cancer, and thus, highly sensitive methods for detecting cancer biomarkers are essential for cancer early diagnosis. Herein, an electrochemical aptamer biosensor based on the 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 the dual recognition of the aptamer to MUC1 and crRNA-Cas12a system to the aptamer. Glucose oxidase (GOD) was loaded on the surface of magnetic Fe3O4@Au (MGNP) via probe single-stranded DNA (pDNA) with the 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-cleaves pDNA and detaches 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 from 1.0 × 10-17 g mL-1 to 1.0 × 10-10 g mL-1. The detection limit was as low as 7.01 × 10-18 g mL-1. The method was applied for the detection of MUC1 in medical samples.
Asunto(s)
Aptámeros de Nucleótidos , Biomarcadores de Tumor , Técnicas Biosensibles , Técnicas Electroquímicas , Glucosa Oxidasa , Mucina-1 , Humanos , Aptámeros de Nucleótidos/química , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Biomarcadores de Tumor/genética , Técnicas Biosensibles/métodos , Proteínas Asociadas a CRISPR/química , Proteínas Asociadas a CRISPR/metabolismo , Sistemas CRISPR-Cas , ADN de Cadena Simple/química , Técnicas Electroquímicas/métodos , Endodesoxirribonucleasas , Enzimas Inmovilizadas/química , Glucosa Oxidasa/química , Glucosa Oxidasa/metabolismo , Oro/química , Peróxido de Hidrógeno/química , Límite de Detección , Nanopartículas de Magnetita/química , Mucina-1/metabolismoRESUMEN
Alkaline phosphatase (ALP) is a major biomarker for clinical diagnosis, but detection methods of ALP are limited in sensitivity and selectivity. In this paper, a novel method for ALP determination is proposed. A photoelectrochemical (PEC) sensor was prepared by growing UiO-tetratopic tetrakis (4-carbox-yphenyl) porphyrin (TCPP) in situ between layered Ti3C2 through a one-pot hydrothermal method. The obtained Schottky heterojunction photoelectric material Ti3C2@UiO-TCPP not only has a large light absorption range but also greatly improves the efficiency of photogenerated electron hole separation and thereby enhances sensitivity for PEC detection. The phosphate group on the phosphorylated polypeptide was utilized to form a Zr-O-P bond with the zirconium ion on UiO-66, and then photocurrent decreases due to the steric hindrance effect of phosphorylated polypeptides, that is, the hindrance of electron transfer between the photoelectric material and a solution. The specific interaction between ALP and phosphorylated polypeptides shears the bond between phosphate and zirconium ion on UiO-66 in the peptides then weakens the hindrance effect and increases the photocurrent, thus realizing ALP detection. The linear range of ALP is 0.03-10,000 U·L-1, and the detection limit is 0.012 U·L-1. The method is highly sensitive and selective, and has been applied in detection of ALP in serum samples.
Asunto(s)
Técnicas Biosensibles , Estructuras Metalorgánicas , Fosfopéptidos , Ácidos Ftálicos , Fosfatasa Alcalina/química , Titanio/química , Circonio/química , Colorantes , Fosfatos , Técnicas Biosensibles/métodos , Límite de Detección , Técnicas Electroquímicas/métodosRESUMEN
A novel extended-gate field-effect transistor (FET) photoelectrochemical (EGFET PEC) sensor was designed for highly sensitive detection of L-cysteine (L-Cys). TiO2 was initially modified on the ITO electrode by the sol-gel dip-coating method and calcined to produce TiO2/ITO. Then, CdS was synthesized on the TiO2 surface by hydrothermal method to obtain the CdS-TiO2 heterojunction material. CdS/TiO2/ITO was connected to the gate of the FET to obtain an EGFET PEC sensor. Under the irradiation of a xenon lamp simulating visible light, the CdS/TiO2 heterojunction composite absorbs light energy to produce photogenerated electron-hole pairs, which have strong photocatalytic oxidation activity and oxidize L-Cys covalently identified by Cd(II) through CdS covalent. These pairs generate a photovoltage that controls the current between the source and the drain to detect L-Cys. Under the optimized experimental conditions, the optical drain current (ID) of the sensor exhibited a good linear relationship with the logarithm of L-Cys in the range of 5.0 × 10-9-1.0 × 10-6 mol/L, and the detection limit was 1.3 × 10-9 mol/L (S/N = 3), which is lower than the values reported by other detection methods. Results showed that the CdS/TiO2/ITO EGFET PEC sensor revealed high sensitivity and good selectivity. The sensor has been used to determine L-Cys in urine samples.