Real time PCR - European Pharmaceutical Review

LAB AUTOMATION ISSUE 2009
Typical images obtained using the Operetta. HeLa cells stained with HOECHST 33342 (blue), Alexa 488 labeled anti-tubulin antibody
and Alexa 647 labeled anti phosphohistone H3 antibody. Control cells (left) and cells treated with 30 μM cytochalasin (right).
‘‘The newly launched
Operetta by Perkin Elmer
provides a complete solution
for new and experienced users
of HCS systems for use in the
life sciences area for studying
and characterising molecular
and morphological events in
cells and tissues in 96 and
384 micro-titre plates’’
HTS on a small selection of
compounds, it is now possible to use
HCS in a much higher capacity and
speed such that in some cases it can
replace altogether the more routinely
used and relatively simple in vitro
biochemical assays. This has been
possible as a result of the fusion of
the outcomes of the advances in
microscopy, image acquisition and
analysis software, computer
processing power, integration into
automated platforms, molecular
biological techniques to construct
tagged target proteins with suitable
labels such as Green Fluorescent
Protein and the development of
fluorescent dyes. HCS screening
offers the possibility of evaluating the
effect of substances (usually small
molecule compounds) on both
phenotypic end-points as well as on
individual cellular events. The newly
launched Operetta by Perkin Elmer
provides a complete solution for new
and experienced users of HCS
systems for use in the life sciences
area for studying and characterising
molecular and morphological events
in cells and tissues in 96 and 384
micro-titre plates. This is also a
valuable tool making it feasible for
more researchers to become involved
in carry out life science research,
target and biomarker identification
and cytotoxicity profiling.
Automated preparation of
assay plates containing
compounds for screening
purposes
Automated compound handling has
usually been treated separately from
the liquid handling of bulk reagents
for assays. Compounds are usually
stored in 100% v/v DMSO as the
solvent for their dissolution in microtitre
plates. Usually, assays will
contain DMSO at concentrations in
the region of 1% v/v, therefore a
100-fold dilution of compound will
need to be carried out. Thus, for an
assay with a volume of 10μl, the
volume of the compound solution
required will be 0.1μl which is usually
added to assay plates first. This is
then followed by the addition of bulk
biological reagents e.g. in the case of
a simple in vitro assay, one addition of
substrate (5μl) and a second addition
of enzyme (5μl) to initiate the
reaction and a third addition (10μl) to
stop the reaction prior to reading the
micro-titre plate. The types of liquid
handling equipment employed to
dispense compounds and bulk
reagents are usually kept separate
as the volumes required are
substantially different.
Numerous technologies have been
developed that are capable of reliable
dispensing of sub-microlitre volumes
of compounds in DMSO whilst
maintaining sufficient throughput for
the production of plates containing
compounds for screening activities.
‘‘The solutions for those
requiring complex high
throughput capabilities
inevitably end up being
financially costly not just in
terms of their initial purchase
price, but also in terms of
preventative maintenance
and service contracts’’
One such example is that based on
capillary action using specially
designed tips for fixed volume
dispensing (Genomic Solutions®
Hummingbird). In this case, as the
same tips are re-used they need to be
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