Issue 39, 2013

An electrochemical assay of polynucleotide kinase activity based on streptavidin–gold nanoparticles and enzymatic amplification

Abstract

An electrochemical method for the determination of polynucleotide kinase (PNK) activity has been proposed with a dual signal amplification strategy. The method relies on the phosphorylation-induced DNA digestion and two-step signal amplification using streptavidin–gold nanoparticle biocomplexes and alkaline phosphatase. In the presence of T4 PNK, the 5′-terminus of the immobilized PNK substrate probe was phosphorylated, and the substrate probe would be cleaved by λ exonuclease after being hybridized with the complementary detection probe biotinylated in the 3′-hydroxyl terminus. As a result, the detection probe would be released, with an electrochemical signal decrease, due to less biotinylated alkaline phosphatase loading on the electrode surface. The electrochemical signal exhibited a linear correlation to the logarithm of PNK concentration ranging from 0.01 U mL−1 to 5 U mL−1 with the detection limit of 0.01 U mL−1. The inhibiting effect of (NH4)2SO4 on the activity of PNK was also evaluated.

Graphical abstract: An electrochemical assay of polynucleotide kinase activity based on streptavidin–gold nanoparticles and enzymatic amplification

Article information

Article type
Paper
Submitted
30 Jun 2013
Accepted
02 Aug 2013
First published
06 Aug 2013

RSC Adv., 2013,3, 18128-18133

An electrochemical assay of polynucleotide kinase activity based on streptavidin–gold nanoparticles and enzymatic amplification

Y. Peng, J. Jiang and R. Yu, RSC Adv., 2013, 3, 18128 DOI: 10.1039/C3RA43315C

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