Biomedical Engineering Reference
In-Depth Information
Fig. 2 Types of variables
which can be examined using
microlitre-scale
chromatography devices
4.3 Microlitre Batch Incubation
4.3.1 Summary of the Technique
Millilitre-scale batch studies in stirred vessels or agitated tubes are used commonly
in early development to examine many variables simultaneously and thus provide
thermodynamic, selectivity and kinetic data [
7
,
37
]. Normally, agitated 96-well
dead-end or filter plates are used, where each well is dosed manually or robotically
with matrix slurries or plaques of known volume. Feed and resin are mixed
together and maintained in suspension for a specified contact time, and the
resulting data can be used to extract information about dynamic column behaviour
[
2
]. Incubation with feed or buffers requires a defined volume per wash and may be
conducted several times if needed to ensure resin saturation or thorough immer-
sion. After incubation, the supernatant is removed, and in dead-end plates, this can
be achieved by gravity or centrifugal sedimentation, followed by careful manual
liquid removal. This method may be time-consuming and risks accidental resin
particle removal; the alternative involves drawing liquid quickly through a filter
plate into a collection plate by centrifugation or under vacuum. Batch plates may
be self- or pre-dispensed, and there are several key factors for successful operation,
including resin and liquid handling, mixing and separation conditions, liquid hold-
up and phase ratios [
7
], as discussed below.
4.3.2 Robotic Slurry Dispensing
Resin is mixed in a reservoir to ensure a homogeneous particle distribution prior to
aliquoting. Resin can be dispensed manually [
20
,
23
], but consistently accurate
resin volumes are more easily achieved robotically, provided that one regulates the
many controlling factors [
7
]; For example, the full resin aliquot must be dispensed
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