Affinity cryogel monoliths for screening for optimal separation conditions and chromatographic separation of cells.
Suitable conditions for separating cells using a chromatographic procedure were evaluated in parallel chromatography on minicolumns. A 96-hole minicolumn plate filled with cryogel monoliths (18.8 mm x 7.1 mm Ø) with immobilized concanavalin A was used. Chromatographic columns (113 mm x 7.1 mm Ø) were used for chromatographic resolution of a mixture of Saccharomyces cerevisiae and Escherichia coli cells. Separation of a cell mixture containing equal amounts of cells of both types performed in a column format under the determined optimal conditions, resulted in a quantitative capture of applied S. cerevisiae cells, while E. coli passed through the column. Bound S. cerevisiae cells were released by flow-induced detachment and by compression of the adsorbent in the presence of 0.3 M methyl alpha-D-manno-pyranoside. The flowthrough and the eluted fractions were analyzed by plate counting and by registering metabolic activity of S. cerevisiae cells in the eluted fractions after capturing on ConA-cryogel monoliths in a 96-minicolumn plate format. The flowthrough fraction contained E. coli cells with nearly 100% purity, whereas the fraction eluted by compression of the adsorbent contained viable S. cerevisiae cells with 95% purity. Thus, an efficient chromatographic separation of cells was achieved using affinity cryogel column.
Dainiak
Maria B
MB
Protista Biotechnology AB, IDEON, SE-223 70 Lund, Sweden.
Galaev
Igor Yu
IY
Mattiasson
Bo
B
D001798:Blood Proteins; D002469:Cell Separation / Q000295:instrumentation / Q000379:methods*; D002846:Chromatography, Affinity / Q000295:instrumentation* / Q000379:methods; D003208:Concanavalin A; D059985:Cryogels; D004926:Escherichia coli / Q000302:isolation & purification*; D005353:Fibronectins; D020100:Hydrogels; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000302:isolation & purification*
D001798:Blood Proteins; D059985:Cryogels; D005353:Fibronectins; D020100:Hydrogels; D003208:Concanavalin A