title: An electrochemical alkaline phosphatase biosensor fabricated with two DNA probes coupled with λ exonuclease.
abstract: In this work we have developed a novel electrochemical biosensor for the detection of alkaline phosphatase (AP) by the use of two complementary DNA probes (DNA 1 and DNA 2) coupled with λ exonuclease (λ exo). Firstly, the 5'-phosphoryl end of DNA 1 is dephosphorylated by AP. Then DNA 1 hybridizes with DNA 2, previously modified on a gold electrode surface. In this double-strand DNA, DNA 2 strand will be promptly cleaved by λ exo with its phosphoryl at the 5' end. After the DNA 2 strand is completely digested, DNA 1 will be released from the double strands and then hybridizes with another DNA 2 strand on the electrode surface, thus the cycle of the release of DNA 1 and the digestion of DNA 2 continues. Since the DNA probes may absorb hexaammineruthenium(III) chloride, the electrochemical species, and the removal of the DNA 2 strand from the electrode surface will result in the decrease of the detected electrochemical signal, which is initially activated by AP, an electrochemical biosensor to assay the activity of AP is proposed in this work. This method may have a linear detection range from 1 to 20 unit/mL with a detection limit of 0.1 unit/mL, and the detection of the enzymatic activity in complex biological fluids can also be realized.
authors.affiliation: Laboratory of Biosensing Technology, School of Life Sciences, Shanghai University, Shanghai, PR China.
authors.forename: Peng
authors.initials: P
authors.lastname: Miao
authors.forename: Limin
authors.initials: L
authors.lastname: Ning
authors.forename: Xiaoxi
authors.initials: X
authors.lastname: Li
authors.forename: Yongqian
authors.initials: Y
authors.lastname: Shu
authors.forename: Genxi
authors.initials: G
authors.lastname: Li
mesh_terms: D000469:Alkaline Phosphatase / Q000032:analysis*; D000818:Animals; D015374:Biosensing Techniques; D001826:Body Fluids / Q000201:enzymology*; D002417:Cattle; D015342:DNA Probes / Q000737:chemistry*; D055664:Electrochemical Techniques; D005090:Exodeoxyribonucleases / Q000737:chemistry*; D057230:Limit of Detection; D017975:Ruthenium Compounds / Q000737:chemistry; D044967:Serum / Q000737:chemistry; D014764:Viral Proteins / Q000737:chemistry*
chemical_list: D015342:DNA Probes; D017975:Ruthenium Compounds; D014764:Viral Proteins; C031584:hexammineruthenium; D005090:Exodeoxyribonucleases; C043586:exo protein, Bacteriophage lambda; D000469:Alkaline Phosphatase