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Novel planar and particle-based microarrays for point-of-care diagnostics
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology.ORCID iD: 0000-0003-4727-6138
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Point-of-care assays are easy-to-use, portable and inexpensive tests that can

be used to aid diagnostics by measuring levels of disease-specific molecules

in settings where access to advanced laboratory equipment and trained

personnel are limited, such as at the patient's bedside or in low resource

parts of developing countries. In order to achieve high multiplexing

capacities, such assays can be based on planar microarrays consisting of

spots immobilized on a flat surface or on particle-based microarrays based

on populations of encoded particles. The aim of the work presented in this

thesis is to develop new point-of-care amenable planar and particle-based

microarrays that allow for highly multiplexed assays while maintaining low

sample-to-result times, complexity and instrumentation requirements.

Paper I demonstrates the use graphically encoded particles for colorimetric

detection of autoantibodies using a consumer-grade flatbed scanner. Four

graphical characters on the surface of each particle allows for millions of

codes and the use of gold nanoparticles as a detection label allows both the

code and the signal intensity to be read out in a single image.

Paper II describes a signal enhancement method that increases the

sensitivity of gold nanoparticle detection on planar microarrays. Using this

method, detection of allergen-specific IgE can be carried out using a

consumer-grade flatbed scanner instead of a more expensive fluorescence

scanner without sacrificing assay performance.

Paper III demonstrates the use of an isothermal DNA amplification method

for detection of adenoviral DNA on a paper-based microarray. Using an

isothermal amplification method eliminates the need for a thermocycler,

reducing the instrumentation required for such detection.

Paper IV shows the use of solid-phase PCR to amplify bacterial DNA directly

on the surface of particles. This strategy reduces assay time by eliminating

the need for separate amplification and hybridisation steps.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2019. , p. 88
Series
TRITA-CBH-FOU ; 2019:13
Keywords [en]
Planar microarrays, Particle-based microarrays, Point-of-care diagnostics, Colorimetric detection, Signal enhancement, Isothermal DNA amplification, Solid-phase DNA amplification
National Category
Medical Biotechnology
Research subject
Biotechnology
Identifiers
URN: urn:nbn:se:kth:diva-244430ISBN: 978-91-7873-101-5 (print)OAI: oai:DiVA.org:kth-244430DiVA, id: diva2:1290486
Public defence
2019-03-22, Science for Life Laboratory, room Air & Fire, Tomtebodavägen 23A, Solna, 10:00 (English)
Opponent
Supervisors
Note

QC 20190221

Available from: 2019-02-21 Created: 2019-02-20 Last updated: 2019-02-21Bibliographically approved
List of papers
1. Towards encoded particles for highly multiplexed colorimetric point of care autoantibody detection
Open this publication in new window or tab >>Towards encoded particles for highly multiplexed colorimetric point of care autoantibody detection
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2017 (English)In: Lab on a Chip, ISSN 1473-0197, E-ISSN 1473-0189, Vol. 17, no 3, p. 549-556Article in journal (Refereed) Published
Abstract [en]

Highly multiplexed point of care tests could improve diagnostic accuracy and differential diagnostic capacity in for instance emergency medicine and low resource environments. Available technology platforms for POC biomarker detection are typically simplex or low-plexed, whereas common lab-based microarray systems allow for the simultaneous detection of thousands of DNA or protein biomarkers. In this study, we demonstrate a novel suspension particle array platform that utilizes 900 mu m bricks for POC amenable colorimetric biomarker detection with an encoding capacity of over two million. Due to the mm-scale size, both the lithographic codes and colorimetric signals of individual particles can be visualized using a consumer grade office flatbed scanner, with a potential for simultaneous imaging of around 19000 particles per scan. The analytical sensitivity of the assay was determined to be 4 ng ml(-1) using an antibody model system. As a proof of concept, autoantibodies toward anoctamin 2 were detected in order to discriminate between multiple sclerosis plasma samples and healthy controls with p < 0.0001 and an inter-assay % CV of 9.44%.

Place, publisher, year, edition, pages
Royal Society of Chemistry, 2017
National Category
Biological Sciences
Identifiers
urn:nbn:se:kth:diva-204746 (URN)10.1039/c6lc01358a (DOI)000395887900019 ()28102419 (PubMedID)2-s2.0-85010943844 (Scopus ID)
Funder
VINNOVAEU, FP7, Seventh Framework Programme, 615458
Note

QC 20170601

Available from: 2017-06-01 Created: 2017-06-01 Last updated: 2019-02-20Bibliographically approved
2. Rapid signal enhancement method for nanoprobe-based biosensing
Open this publication in new window or tab >>Rapid signal enhancement method for nanoprobe-based biosensing
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2017 (English)In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, article id 6837Article in journal (Refereed) Published
Abstract [en]

The introduction of nanomaterials as detection reagents has enabled improved sensitivity and facilitated detection in a variety of bioanalytical assays. However, high nanoprobe densities are typically needed for colorimetric detection and to circumvent this limitation several enhancement protocols have been reported. Nevertheless, there is currently a lack of universal, enzyme-free and versatile methods that can be readily applied to existing as well as new biosensing strategies. The novel method presented here is shown to enhance the signal of gold nanoparticles enabling visual detection of a spot containing < 10 nanoparticles. Detection of Protein G on paper arrays was improved by a 100-fold amplification factor in under five minutes of assay time, using IgG-labelled gold, silver, silica and iron oxide nanoprobes. Furthermore, we show that the presented protocol can be applied to a commercial allergen microarray assay, ImmunoCAP ISAC sIgE 112, attaining a good agreement with fluorescent detection when analysing human clinical samples.

Place, publisher, year, edition, pages
NATURE PUBLISHING GROUP, 2017
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-212341 (URN)10.1038/s41598-017-07030-0 (DOI)000406610000088 ()2-s2.0-85026428495 (Scopus ID)
Note

QC 20170823

Available from: 2017-08-23 Created: 2017-08-23 Last updated: 2019-02-20Bibliographically approved
3. Adenoviral detection by recombinase polymerase amplification and vertical flow paper microarray.
Open this publication in new window or tab >>Adenoviral detection by recombinase polymerase amplification and vertical flow paper microarray.
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2019 (English)In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 411, no 4, p. 813-822Article in journal (Refereed) Published
Abstract [en]

Respiratory viral infections often mimic the symptoms of infections caused by bacteria; however, restricted and targeted administration of antibiotics is needed to combat growing antimicrobial resistance. This is particularly relevant in low-income settings. In this work, we describe the use of isothermal amplification of viral DNA at 37 °C coupled to a paper-based vertical flow microarray (VFM) setup that utilizes a colorimetric detection of amplicons using functionalized gold nanoparticles. Two oligonucleotide probes, one in-house designed and one known adenoviral probe were tested and validated for microarray detection down to 50 nM using a synthetic target template. Furthermore, primers were shown to function in a recombinase polymerase amplification reaction using both synthetic template and viral DNA. As a proof-of-concept, we demonstrate adenoviral detection with four different adenoviral species associated with respiratory infections using the paper-based VFM format. The presented assay was validated with selected adenoviral species using the in-house probe, enabling detection at 1 ng of starting material with intra- and inter-assay %CV of ≤ 9% and ≤ 13%. Finally, we validate our overall method using clinical samples. Based on the results, the combination of recombinase polymerase amplification, paper microarray analysis, and nanoparticle-based colorimetric detection could thus be a useful strategy towards rapid and affordable multiplexed viral diagnostics.

Place, publisher, year, edition, pages
Springer, 2019
Keywords
Adenoviral, RPA, VFM
National Category
Medical Biotechnology
Identifiers
urn:nbn:se:kth:diva-244426 (URN)10.1007/s00216-018-1503-y (DOI)000456132900003 ()30498984 (PubMedID)2-s2.0-8505759966 (Scopus ID)
Note

QC 20190222

Available from: 2019-02-20 Created: 2019-02-20 Last updated: 2019-02-22Bibliographically approved
4. Solid-phase PCR on graphically encoded microparticles for multiplexed colorimetric detection of bacterial DNA
Open this publication in new window or tab >>Solid-phase PCR on graphically encoded microparticles for multiplexed colorimetric detection of bacterial DNA
Show others...
(English)Manuscript (preprint) (Other academic)
National Category
Medical Biotechnology
Research subject
Biotechnology
Identifiers
urn:nbn:se:kth:diva-244428 (URN)
Note

QC 20190221

Available from: 2019-02-20 Created: 2019-02-20 Last updated: 2019-02-21Bibliographically approved

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