Macroporous monolithic hydrogels in a 96-minicolumn plate format for cell surface-analysis and integrated binding/quantification of cells

Abstract

Macroporous monolithic hydrogels (cryogel monoliths; rods 12.5 mm × 7.1 mm diameter) are elastic, sponge-like materials with large (10–100 μm), interconnected pores. Phenyl- and IMAC-(Me(II)-iminodiacetic acid)-cryogel monoliths were inserted into the open-ended wells of a standard 96-well plate, forming a system of 96 drainage-protected minicolumns, and were used in a parallel assay of hydrophobicity and affinity to immobilized metal ions of wild type Escherichia coli cells , recombinant E. coli cells with poly-His peptide displayed on the cell surface, and Bacillus halodurans cells in different growth phases. Bound cells were eluted with standard eluents or were detached by mechanical compression of affinity cryogel monoliths in the case of strongly bound cells. The possibility to carry out high throughput viability assays of bound cells was demonstrated on an example of analysis of recombinant E. coli cells bound to Cu(II)-IDA-cryogel monoliths and of yeast cells bound to ConA-cryogel monoliths, where the metabolic activity of cells was measured using tetrazolium salt XTT and pH indicator, neutral red, respectively. The developed system can be used for the rapid optimization of chromatographic separation of cells and for detection of cells of interest from a large number of medical and food samples.