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AssigneeBruker Daltonik GmbH
InventorsChristian Neususs, Matthias Pelzing, Jochen Franzen
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The invention relates to methods used for mass spectrometric analysis of substances separated by capillary electrophoresis, in particular biopolymers such as proteins, proteoglycanes or other protein conjugates or their digest peptides. The invention consists in reducing the electrophoresis voltage upon appearance of the first analytically interesting substance, thereby maintaining the high separation power and gaining sensitivity. With direct coupling, e.g. by electrospray, the electrophoretic voltage may be directly controlled by the ion current measured by the mass spectrometer.
Method for coupling electrophoresis in capillaries or microchannels for the separation of substances in a mixture to mass spectrometric analysis of the thus separated substances, where certain groups of substances are of particular interest,
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Details; Accessories: arrangements for investigating the separated zones, e.g. localising zones
Investigating or analysing materials by determining their chemical or physical properties
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The invention relates to methods used for mass spectrometric analysis of substances separated by capillary electrophoresis, in particular biopolymers such as proteins, proteoglycanes or other protein conjugates or their digest peptides.
In the analysis of biochemical polymers, in particular proteomics, separation methods with the highest possible resolution are sought in order to find alternatives to the difficult to perform 2D gel electrophoresis method. The methods mainly considered are those where mixtures of proteins or their conjugates are digested before separation to produce hundreds, if not thousands, of digest peptides which are then separated from each other and subjected to mass spectrometric analysis. The analysis is not restricted to the determination of their molecular weights but also of their structures by measuring the daughter-ion spectra of fragmented ions.
Capillary clectrophoresis (CE) has been found to be the separation method with the highest separation efficiency so far. However, the separation efficiency is usually reduced significantly by the coupling methods (CE/MS) with mass spectrometers (MS). This applies to both ionization methods for the mass spectrometers: electrospray ionization (ESI) and matrix assisted laser desorption and ionization (MALDI).
FIG. 1 shows a typical electrophorogram for a mixture of digest peptides. The appearance of the digest peptides starts after 6 minutes and ends after 10 minutes. After 15 minutes, some neutral substances appear which have been flushed out with the electroosmotic flow (EOF). These substances are not digest peptides.
FIG. 2 shows three electrophorograms for a mixture of peptides using a 55 cm×50 m×360 m capillary (length×internal diameter×external diameter):
(A) an electrophorogram at 30 kV when the invention has not been used,
The favorable embodiments for ionization by electrospray (ESI) will be given first, followed by those for ionization by matrix supported laser desorption (MALDI).
The effect of the invention is well shown in FIG. 2, exhibiting three electrophorograms for a mixture of peptides using a 55 cm×50 m×360 m capillary (length×internal diameter×external diameter) in connection with electrospray ionization:
(A) an electrophorogram at 30 kV when the invention has not been used,