Change search
ReferencesLink to record
Permanent link

Direct link
Miniaturized Techniques for Protein Analysis
KTH, Superseded Departments, Chemistry.
2004 (English)Doctoral thesis, comprehensive summary (Other scientific)
Abstract [en]

Proteins are a highly diversified group of molecules, andfor their study, advanced analytical tools are required. Inparticular, a need for high-throughput techniques has emergedin order to enable the characterization of large sets ofproteins. In this thesis, improved techniques for proteinseparations as well as new tools for the mass spectrometricanalysis of proteins are described.

In the work, presented in the first part of the thesis, arefined extract containing proteases from Antarctic krill (Euphausia superba) was separated and characterized bymeans of capillary electrophoresis (CE) and mass spectrometry(MS). Tailored CE separations of the krill extract revealed thepresence of approximately 50 components. In addition, adetailed CE and MS analysis of fractions, containing individualkrill proteases has been carried out. Trypsin-like proteasesfrom krill exhibited a 12-fold and a 60-fold higher digestionefficiency at 37 °C and 2 °C respectively compared todigests performed with bovine trypsin. Furthermore, thecleavage specificity of the trypsin-like proteases wasstudied.

In the last part of the thesis, novel concepts forchip-based nanoelectrospray (nanoESI) and matrix-assisted laserdesorption/ionization (MALDI) mass spectrometry are described.First, a micromachined silicon chip with a two-dimensionalmatrix of out-ofplane nanoESI needles for high-throughputanalysis was fabricated. A two-fold improvement insignal-to-noise reproducibility was obtained. Second, achip-based target for MALDI was developed, which featured pairsof elevated 50×50 µm anchors in close proximity. Theanchors were individually addressable with sample solution. Theminiaturized sample preparations at close distance to eachother allowed a simultaneous ionization of a physicallyseparated sample and standard by one single laser pulse. Thisresulted in a twofold reduction of relative mass errors.Moreover, ion suppression of the analyte was significantlyreduced. The effective utilization of the sample resulted in adetection limit of ca 200 zeptomole of angiotensin I.

Key words:Proteins, peptides, proteases, Antarctickrill,Euphausia superba, capillary electrophoresis,fluorosurfactants, mass spectrometry, nanoelectrospray, ESI,MALDI, chip, high-throughput, reproducibility, sensitivity andmass accuracy.

Place, publisher, year, edition, pages
Stockholm: Kemi , 2004. , v, 74 p.
Keyword [en]
Proteins, peptides, proteases, Antarctic krill, Euphausia superba, capillary electrophoresis, fluorosurfactants, mass spectrometry, nanoelectrospray, ESI, MALDI, chip, high-throughput, reproducibility, sensitivity, mass accuracy
URN: urn:nbn:se:kth:diva-3802ISBN: 91-7283-783-7OAI: diva2:9656
Public defence
2004-06-11, 00:00
Available from: 2004-06-10 Created: 2004-06-10 Last updated: 2012-03-21

Open Access in DiVA

fulltext(2275 kB)729 downloads
File information
File name FULLTEXT01.pdfFile size 2275 kBChecksum SHA-1
Type fulltextMimetype application/pdf

Search in DiVA

By author/editor
Sjödahl, Johan
By organisation

Search outside of DiVA

GoogleGoogle Scholar
Total: 729 downloads
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Total: 388 hits
ReferencesLink to record
Permanent link

Direct link