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How to use BioTrap 500
A conventional 6-port switching valve is used to connect
BioTrap 500 (the extraction column) with the analytical column
(see figures below). By using a 6-port valve with an electric
actuator and injecting the samples by an autosampler, it is
possible to obtain a complete automatization of the system. The
macromolecular material is separated from low molecular weight
material on the extraction column. The procedure is described in
detail in the text and the figures below. The
extraction mobile phase is a buffer with low content of
organic modifier, optimized in order to get high recoveries of
the analyte and an effective rinsing of the extraction column. The analytical mobile phase is used for
transfer of the analytes from the extraction column to the
analytical column, and is optimized for the final separation on
the analytical column.
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- Extraction position - sample injection and
separation of the analyte(s) from high molecular
weight compounds (e.g. proteins). In this
position the sample (e.g. plasma) is injected
onto BioTrap 500 (the extraction column). The
plasma proteins and other macromolecular
compounds will be excluded from the pores and
eluted to waste. Low molecular weight compounds
penetrate the pores of the particles and are
retained by the hydrophobic groups (C18/C8) on
the inner surface of the particles. When the
proteins have been washed out from the column,
the valve can be switched to the elution
position. To extend the lifetime of BioTrap 500,
the samples should be centrifuged prior to
injection.
- Elution position - transfer of the adsorbed
compounds from the extraction column to the
analytical column. In this position the
analytes will be eluted from the extraction
column onto the analytical column by the
analytical mobile phase. When the analytes have
been transferred to the analytical column, the
valve can be switched back to the extraction
position.
- Extraction position - the final separation of
the analytes on the analytical column and
reequilibration of the extraction column. In
this position the final separation of the
analytes is achived on the analytical column.
Simultaneously, the extraction column is
reequilibrated with the extraction mobile phase
and the system will be ready for the next
injection.
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If you have any questions regarding the above,
don´t hesitate to contact
ChromTech!
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