Premium
Capillary isoelectric focusing of proteins utilizing poly(vinylpyrrolidone)‐ and plexiglas‐coated columns
Author(s) -
Palm Anders,
ZaragozaSundqvist Maximiliano,
MarkoVarga György
Publication year - 2004
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.200301596
Subject(s) - isoelectric focusing , coating , chromatography , chemistry , reproducibility , silane , capillary action , analytical chemistry (journal) , materials science , composite material , organic chemistry , enzyme
Fused‐silica capillaries chemically derivatized with silane/poly(vinylpyrrolidone) (PVP) or dynamically modified with plexiglas [poly(methyl methacrylate)] were prepared and evaluated with regard to column stability and separation performance for capillary isoelectric focusing of standard proteins. The PVP coating showed the better stability and was good for at least 100 runs while the plexiglas coating started to deteriorate after about 30 runs. The time spent for the plexiglas coating is about 40 minutes while the PVP coating requires two days. The migration time reproducibility was better with the PVP capillary (RSD 0.7–1.6%, n = 5) compared to the plexiglas‐coated column (RSD 1.2–2.9%, n = 5) while peak area and height varied over a similar interval (RSD 2–28.1% area; 0.9–22.7% height, n = 5). The two most consistent proteins in this evaluation, viz. myoglobin A and carbonic anhydrase II, showed linear dynamic ranges between 5–150 and 5–50 μg/mL, and limits of detections at 2 and 1 μg/mL, respectively, employing UV detection at 280 nm.