期刊
ACTA BIOCHIMICA POLONICA
卷 66, 期 3, 页码 299-304出版社
ACTA BIOCHIMICA POLONICA
DOI: 10.18388/abp.2019_2774
关键词
RNA biological samples; redox-active layer; electrochemical genosensor; square wave voltammetry; mismatched base pairs
资金
- National Centre for Research and Development (NCBiR) [PBS2/A7/14/2014, 679/N-BELGIA/2010/0]
- COST Action [CM10005]
- Institute of Animal Reproduction and Food Research, Polish Academy of Sciences, Olsztyn, Poland
An electrochemical genosensor based on an epoxyphenanthroline-Fe(III)-NH2-ssDNA layer for the detection of RNA derived from Avian Influenza is presented. The biosensor preparation consists of: (I) modification of gold electrodes with aminoethanethiol, (II) modification of the self-assembled monolayer of aminoethanethiol with 5,6-epoxy-5,6-dihydro-[1,10]-phenanthroline using click chemistry, (Ill) a first step of complexation of Fe(III) by 5,6-epoxy-5,6-dihydro-[1,10]-phenanthroline, (IV) a second step of complexation of Fe(III) by 5,6-epoxy-5,6-dihydro-[1,10]-phenanthroline, (V) immobilization of the single stranded amino-DNA probe via click chemistry between epoxy and amino groups. The interactions between the ssDNA probe and RNA targets were explored with Osteryoung Square Wave Voltammetry. The genosensor showed a remarkable detection limit of 3 copies/mu L (5 aM) for RNA extracted from A/swan/Poland/305/06 (H5N1) containing a fully complementary sequence. A linear dynamic range for this sequence was observed from 3.0x10(3) to 3.0x10(5) [copies/mu l]. RNA extracted from A/mallard/Poland/446/09 (H7N7), containing a non-complementary sequence, generated a much weaker response. Moreover, the developed genosensor allows to distinguish RNA present in biological samples having 2, 3 and 4 mismatches. This biosensing approach can become a potential alternative tool for detecting RNA samples in biomedical research and early clinical diagnosis of avian influenza viruses.
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