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One-step scalable fluorescent microgel bioassay for endogenous viral miR-US4-5p ultrasensitive detection

Abstract

Human Cytomegalovirus (hCMV) infection is the leading cause of new birth defects and death in immunosuppressed people. Traditional techniques require long time and costly analyses, sometimes cause of false positive results, thus a rapid and accurate detection is necessary. Recently, hcmv-miR-US4-5p has been selected as biomarker for cytomegalovirus diagnosis and follow-up. Herein, we propose a bioassay based on microgels endowed by optical fluorescent oligonucleotide probes for the detection of circulating endogenous hcmv-microRNAs. In particular, a double strand probe, based on fluorescence recovery after target capture, is conjugated on microgels and the probe density is opportunely optimised. Then the microgels are directly mixed with the sample. The fluorescence read-out is measured as function of target concentration at fixed number of microgel per tube. As beads-based assay the performances of optical detection in terms of dynamic working range and limit of detection can be finely tuned by playing with the number of microgels per tube. The limit of detection of the assay can be tuned in the range from 39.1 fM to 156 aM by changing the microgel concentration respectively from 50 µg/mL to 0.5 µg/mL. The assay results specific for the selected target, stable over a one-year time span and it is not affected by the presence of human serum. Therefore, the bioassay based on microgel might represent a flexible platform able to predict, identify and follow-up several diseases, monitoring free circulating oligonucleotide in body fluids.

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Publication details

The article was received on 09 Nov 2018, accepted on 05 Dec 2018 and first published on 06 Dec 2018


Article type: Paper
DOI: 10.1039/C8AN02166J
Citation: Analyst, 2018, Accepted Manuscript
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    One-step scalable fluorescent microgel bioassay for endogenous viral miR-US4-5p ultrasensitive detection

    T. M. Caputo, A. Cummaro, V. Lettera, A. Mazzarotta, E. Battista, P. A. Netti and F. Causa, Analyst, 2018, Accepted Manuscript , DOI: 10.1039/C8AN02166J

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