Preparative isoelectric focusing of microorganisms in cellulose-based separation medium and subsequent analysis by CIEF and MALDI-TOF MS

- Preparative IEF in cellulose-based medium was used for separation of cells.
- The cells of M. luteus, Dietzia sp. and R. mucilaginosa strains were examined.
- The pI's of these colored microorganisms were determined by CIEF.
- The collected fractions of cells were analyzed by CIEF, MALDI or cultivated.

Pre-separation and pre-concentration of bacteria is an important step especially when they are uncultured and bacterial concentration in the matrix is low. This study describes a preparative method based on isoelectric focusing of colored microorganisms in a cellulose-based separation medium from a high conductivity matrix. The isoelectric points found for the examined cells were 1.8 for Micrococcus luteus, 3.5 for Dietzia sp., and 4.7 for Rhodotorula mucilaginosa using capillary isoelectric focusing. The final positions of the zones of colored microbial cells in the cellulose-bed are indicated by colored pI markers. Segments of the separation medium with cells were harvested by a spatula, simply purified using centrifugation and analyzed by capillary isoelectric focusing and matrix-assisted laser desorption/ionization time of flight mass spectrometry. The determined recovery ranged from 78% to 93%. The viability of the harvested cells was verified by their cultivation.

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