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Biosensor based protein profiling on reverse phase serum microarray
KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
KTH, School of Biotechnology (BIO), Proteomics (closed 20130101). KTH, Centres, Science for Life Laboratory, SciLifeLab.
2012 (English)In: Journal of Proteomics & Bioinformatics, ISSN 0974-276X, Vol. 5, no 8, 185-189 p.Article in journal (Refereed) Published
Abstract [en]

The reverse phase serum microarray format enables multi-parallel and simultaneous analysis of literally thousands of samples, a feature which is of uttermost importance for protein profiling of clinical samples. We have here screened 2400 serum samples for their potential IgA deficiency by using a fluorescence based reverse phase serum microarray platform and a biosensor based label-free microarray platform for verification and also compared our microarray-results to clinical routine ELISA. We have been able to identify possible IgA-deficiencies and to show the suitability of our microarray-platforms for large-scale screening of clinical serum samples. The two microarray methods show reproducibility and correlation towards each other and low variation between replicates within each platform. Both of the microarray platforms show less agreement towards ELISA. The fluorescence based microarray method has been shown to be applicable for large-scale screening of clinically important serum samples for detection of possibly IgA-deficient patients. Furthermore, it was found that the microarray based biosensor method could be used for determining the relative differences in concentration of IgA between the samples.

Place, publisher, year, edition, pages
2012. Vol. 5, no 8, 185-189 p.
Keyword [en]
Biosensor, Label-free, Protein profiling and screening, Reverse phase serum microarray, SPR
National Category
Biological Sciences
URN: urn:nbn:se:kth:diva-103635DOI: 10.4172/jpb.1000233ScopusID: 2-s2.0-84866123312OAI: diva2:561208
Science for Life Laboratory - a national resource center for high-throughput molecular bioscience

QC 20121017

Available from: 2012-10-17 Created: 2012-10-17 Last updated: 2015-05-28Bibliographically approved
In thesis
1. On Generation and Applications of High-Density Protein Microarrays
Open this publication in new window or tab >>On Generation and Applications of High-Density Protein Microarrays
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Affinity proteomics has experienced rapid development over the last two decades and one of the most promising platforms to emerge are the protein microarrays. The combination of affinity reagents and miniaturisation enables assays for simultaneous high throughput and sensitive protein analysis. Due to the combination of these desrable properties, a multitude of protein array platforms for rapid and efficient study of proteomes and protein interactions are in use today. Although the protein microarray field has more than two decades of history to look back on the development of new protein microarray platforms continues to this day and beyond.

In the paper I in this thesis, a microarray of eluates from dried blood spot samples collected from neonates were designed and utilised for detection of complement factor 3 (C3) deficiency. The data acquired from the microarrays platform were compared to C3 levels obtained through enzyme-linked immunosorbant assay (ELISA), and the microarray assay were found to separate the C3 deficient samples from the controls. The conclusion of this investigation was that the microarray platform would be suitable for high-throughput screening of C3 deficiency in neonates. Paper II outlines the work in developing a multiplex platform for validation of affinity reagents. A set of 398 affinity binders, originating from five research groups, were profiled against 432 antigens and representing both polyclonal rabbit antibodies, monoclonal mouse antibodies, and recombinant single-chain variable fragments. Approximately 50% of the binders were found to preferably recognise their intended target while 10% of the binders did not generate any, or low, signals with their respective targets. For paper III, a reverse phase array (RPPA) platform using fluorescence-based detection of IgA deficiency in over 2.000 samples where validated on a label-free detection system and ELISA. The data from the label-free platform and the RPPA were found agree well with each other while data from ELISA did with neither of them. It was found that the label-free platform proved to be well-suited for detection of IgA in serum. Paper IV describes one of the world’s largest protein microarrays containing 21.120 recombinant protein fragments. We describe some of the possible applications of these large-scale arrays, such as binding profiles for the validation of antibodies with 11.520 and 21.120 recombinant proteins, as well as screening for autoimmunity in human serum samples.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2015. ix, 63 p.
TRITA-BIO-Report, ISSN 1654-2312 ; 2015:12
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
urn:nbn:se:kth:diva-168165 (URN)978-91-7595-579-7 (ISBN)
Public defence
2015-06-12, Inghe-salen, Tomtebodavägen 18a, Solna, 10:00 (English)

QC 20150528

Available from: 2015-05-28 Created: 2015-05-27 Last updated: 2016-01-28Bibliographically approved

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