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Electrokinetic sandwich assay and DNA mediated charge amplification for enhanced sensitivity and specificity
Uppsala Univ, Dept Elect Engn, Angstrom Lab, Uppsala, Sweden..ORCID-id: 0000-0002-2794-9158
KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap, Proteinvetenskap.ORCID-id: 0000-0002-6552-8426
KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap.
KTH, Skolan för kemi, bioteknologi och hälsa (CBH), Proteinvetenskap.ORCID-id: 0000-0001-7755-2661
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2021 (Engelska)Ingår i: Biosensors & bioelectronics, ISSN 0956-5663, E-ISSN 1873-4235, Vol. 176, artikel-id 112917Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

An electrical immuno-sandwich assay utilizing an electrokinetic-based streaming current method for signal transduction is proposed. The method records the changes in streaming current, first when a target molecule binds to the capture probes immobilized on the inner surface of a silica micro-capillary, and then when the detection probes interact with the bound target molecules on the surface. The difference in signals in these two steps constitute the response of the assay, which offers better target selectivity and a linear concentration dependent response for a target concentration within the range 0.2-100 nM. The proof of concept is demonstrated by detecting different concentrations of Immunoglobulin G (IgG) in both phosphate buffered saline (PBS) and spiked in E. coli cell lysate. A superior target specificity for the sandwich assay compared to the corresponding direct assay is demonstrated along with a limit of detection of 90 pM in PBS. The prospect of improving the detection sensitivity was theoretically analysed, which indicated that the charge contrast between the target and the detection probe plays a crucial role in determining the signal. This aspect was then experimentally validated by modulating the zeta potential of the detection probe by conjugating negatively charged DNA oligonucleotides. The length of the conjugated DNA was varied from 5 to 30 nucleotides, altering the zeta potential of the detection probe from -9.3 +/- 0.8 mV to -20.1 +/- 0.9 mV. The measurements showed a clear and consistent enhancement of detection signal as a function of DNA lengths. The results presented here conclusively demonstrate the role of electric charge in detection sensitivity as well as the prospect for further improvement. The study therefore is a step forward in developing highly selective and sensitive electrokinetic assays for possible application in clinical investigations.

Ort, förlag, år, upplaga, sidor
Elsevier BV , 2021. Vol. 176, artikel-id 112917
Nyckelord [en]
Sandwich assay, Label-free detection, DNA-Conjugated affinity probes, Biosensor, Electrokinetics, Streaming current, Zeta potential, Improved specificity and sensitivity
Nationell ämneskategori
Biokemi och molekylärbiologi
Identifikatorer
URN: urn:nbn:se:kth:diva-291976DOI: 10.1016/j.bios.2020.112917ISI: 000612677000002PubMedID: 33421763Scopus ID: 2-s2.0-85098774737OAI: oai:DiVA.org:kth-291976DiVA, id: diva2:1541335
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QC 20210331

Tillgänglig från: 2021-03-31 Skapad: 2021-03-31 Senast uppdaterad: 2022-06-25Bibliografiskt granskad

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Sahu, Siddharth SourabhStiller, ChristianePaz Gomero, ElizabethNagy, AbelEriksson Karlström, AmelieLinnros, JanDev, Apurba
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ProteinvetenskapProteinvetenskapAlbanova VinnExcellence Center for Protein Technology, ProNovaFotonik
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