BMC Biotechnology | |
Identification of proteins in laser-microdissected small cell numbers by SELDI-TOF and Tandem MS | |
Grazyna Kwapiszewska1  Markus Meyer2  Ralf Bogumil2  Rainer M Bohle1  Werner Seeger1  Norbert Weissmann3  Ludger Fink1  | |
[1] Department of Pathology, Justus-Liebig University Giessen, Langhansstr.10, 35392 Giessen, Germany | |
[2] Ciphergen Biosystems GmbH, Göttingen, Germany | |
[3] Department of Internal Medicine II, Justus-Liebig University Giessen, 35392 Giessen, Germany | |
Others : 1154462 DOI : 10.1186/1472-6750-4-30 |
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received in 2004-07-13, accepted in 2004-12-03, 发布年份 2004 | |
【 摘 要 】
Background
Laser microdissection allows precise isolation of specific cell types and compartments from complex tissues. To analyse proteins from small cell numbers, we combine laser-microdissection and manipulation (LMM) with mass spectrometry techniques.
Results
Hemalaun stained mouse lung sections were used to isolate 500–2,000 cells, enough material for complex protein profiles by SELDI-TOF MS (surface enhanced laser desorption and ionization/time of flight mass spectrometry), employing different chromatographic ProteinChip® Arrays. Initially, to establish the principle, we identified specific protein peaks from 20,000 laser-microdissected cells, combining column chromatography, SDS-PAGE, tryptic digestion, SELDI technology and Tandem MS/MS using a ProteinChip® Tandem MS Interface. Secondly, our aim was to reduce the labour requirements of microdissecting several thousand cells. Therefore, we first defined target proteins in a few microdissected cells, then recovered in whole tissue section homogenates from the same lung and applied to these analytical techniques. Both approaches resulted in a successful identification of the selected peaks.
Conclusion
Laser-microdissection may thus be combined with SELDI-TOF MS for generation of protein marker profiles in a cell-type- or compartment-specific manner in complex tissues, linked with mass fingerprinting and peptide sequencing by Tandem MS/MS for definite characterization.
【 授权许可】
2004 Kwapiszewska et al; licensee BioMed Central Ltd.
【 预 览 】
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20150407104553448.pdf | 842KB | download | |
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Figure 2. | 45KB | Image | download |
Figure 1. | 174KB | Image | download |
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