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Miniaturized and Au...
Miniaturized and Automated High-Throughput Verification of Proteins in the ISET Platform with MALDI MS
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- Adler, Belinda (author)
- Lund University,Lunds universitet,Avdelningen för Biomedicinsk teknik,Institutionen för biomedicinsk teknik,Institutioner vid LTH,Lunds Tekniska Högskola,Department of Biomedical Engineering,Departments at LTH,Faculty of Engineering, LTH
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- Boström, Tove (author)
- KTH,Proteomik
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- Ekström, Simon (author)
- Lund University,Lunds universitet,Avdelningen för Biomedicinsk teknik,Institutionen för biomedicinsk teknik,Institutioner vid LTH,Lunds Tekniska Högskola,Department of Biomedical Engineering,Departments at LTH,Faculty of Engineering, LTH
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- Hober, Sophia (author)
- KTH,Proteomik
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- Laurell, Thomas (author)
- Lund University,Lunds universitet,Avdelningen för Biomedicinsk teknik,Institutionen för biomedicinsk teknik,Institutioner vid LTH,Lunds Tekniska Högskola,Department of Biomedical Engineering,Departments at LTH,Faculty of Engineering, LTH
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(creator_code:org_t)
- 2012-09-24
- 2012
- English.
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In: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 84:20, s. 8663-8669
- Related links:
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http://dx.doi.org/10...
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https://urn.kb.se/re...
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https://doi.org/10.1...
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https://lup.lub.lu.s...
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Abstract
Subject headings
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- A major bottleneck in high-throughput protein production is the validation step, which is why parallel and automated sample processing methods are highly desirable. Also, a miniaturized sample preparation format is preferred, as the reduction of reagent volumes significantly decreases the analysis cost per sample. We have developed an automated and miniaturized protein sequence verification protocol for recombinant proteins utilizing peptide mass fingerprinting and MS/MS analysis. The integrated selective enrichment target (ISET) platform, previously developed in our group, with its dual functionality, being both a sample preparation platform and a MALDI target plate, is employed. All steps including immobilized metal ion affinity chromatography of protein on cobalt-loaded beads, tryptic digestion, and MALDI MS analysis are performed in an array format, without any sample transfers, on the same ISET chip. The automated configuration reduced the sample preparation time significantly. Starting with crude lysate, a full plate of 48 purified, digested samples prepared for MALDI-MS can be generated in 4 h, with only 30 min of operator involvement. This paper demonstrates the utility of the method by parallel analysis of 45 His-tagged human recombinant proteins.
Subject headings
- NATURVETENSKAP -- Kemi -- Analytisk kemi (hsv//swe)
- NATURAL SCIENCES -- Chemical Sciences -- Analytical Chemistry (hsv//eng)
Keyword
- Assisted-Laser-Desorption/Ionization
- Selective Enrichment Target
- Spectrometry-Based Proteomics
- Mass-Spectrometry
- Sample Preparation
- Affinity-Chromatography
- Recombinant Proteins
- Microfluidic Chips
- Identification
- Expression
Publication and Content Type
- ref (subject category)
- art (subject category)
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