Utilization of newly developed immobilized enzyme reactors for preparation and study of immunoglobulin G fragments
Language English Country Netherlands Media print
Document type Journal Article, Research Support, Non-U.S. Gov't
PubMed
15236681
DOI
10.1016/j.jchromb.2004.04.035
PII: S1570023204003988
Knihovny.cz E-resources
- MeSH
- Bioreactors * MeSH
- Electrophoresis, Polyacrylamide Gel MeSH
- Enzymes metabolism MeSH
- Mass Spectrometry MeSH
- Immunoglobulin G metabolism MeSH
- Immunoglobulin Fragments chemistry metabolism MeSH
- Molecular Sequence Data MeSH
- Amino Acid Sequence MeSH
- Spectroscopy, Fourier Transform Infrared MeSH
- Chromatography, High Pressure Liquid MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Enzymes MeSH
- Immunoglobulin G MeSH
- Immunoglobulin Fragments MeSH
The newly developed immobilized enzyme reactors (IMERs) with proteolytic enzymes chymotrypsin, trypsin or papain were used for specific fragmentation of high molecular-mass and heterogeneous glycoproteins immunoglobulin G (IgG) and crystallizable fragment of IgG (Fc). The efficiency of splitting or digestion were controlled by RP-HPLC. The specificity of digestion by trypsin reactor was controlled by MS. IMERs (trypsin immobilized on magnetic microparticles focused in a channel of magnetically active microfluidic device) was used for digestion of the whole IgG molecule. The sufficient conditions for IgG digestion in microfluidic device (flow rate, ratio S:E, pH, temperature) were optimized. It was confirmed that the combination of IMERs with microfluidic device enables efficient digestion of highly heterogeneous glycoproteins such as IgG in extremely short time and minimal reaction volume.
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