Assay and Drug Development Technologies - 10

GIULIANO ET AL.
Figure 2 demonstrates how the enormous amount ofdata from
one of these general BioApplications can be easily visualized
with the appropriate data viewing software. Finally, the introduction
of products like Cellomics vHCS: Build, permits
users to augment the BioApplication logic to create additional
features to interpret results. This automated approach now
yields both flexibility and specificity in simple solutions.
With newly available specific and general BioApplications,
the true power of HCS is realized. These BioApplications can
measure and correlatemultiple features ofall cells imaged and
report results at the cell and population levels. This enables
higher throughput, sophisticated assays, and deeper biological
information. The ability to distinguish, measure, and
compare responses between subpopulations of cells enables
detection of cellular events not achievable with assays reporting
only single values for a well. Furthermore, multiplexing
several of these advanced BioApplications during a
screening campaign builds more layers of biological information
from which even better decisions can be made.
BioApplication software modules are one of the most critical
components of an integrated HCS platform. BioApplications
are essentially software ''experts'' that translate image
data into biological information within the proper context. It
will soon be possible to use these powerful processing and
analysis modules on images and data sets acquired from a
variety of sources, including research epifluorescence and
confocal microscopes, laser scanning-cytometry systems, and
other HCS instrumentation.
REAGENTS
The selection and application of the optimal combination of
fluorescent probes tomeasure multiple, specific cellular features
are key to successful HCS whether it is done in fixed-end-point
or kinetic mode. While distinct classes of fluorescent probes are
used for fixed-end-point or live cell kinetic assays, there is often
functional overlap between the probes. For example, fluorescent
protein biosensors are, in several cases, used in fixed-end-point
HCS assays even though they are designed to measure both the
temporal and spatial components of specific cellular processes.44
For HCS assays performed in the fixed-end-point and kinetics
modes, Cellomics has produced HCS reagent kits. These
kits are comprised of a validated set of fluorescent and nonfluorescent
reagents, protocols for sample preparation that include
automated and manual approaches, instruction on how
the appropriate HCSReader is set up to extract information from
the samples, and cell-based data that demonstrate the performance
of the kit in a real-world screening environment.
The true power of HCS resides in its capacity to integrate
signals from multiple reagents within populations of single
10 ASSAY and Drug Development Technologies
cells. Nevertheless, taking advantage of this particular
strength means that multiparameter reagent designs can become
extremely complex, especially if the assay contains up
to six channels of fluorescence-based information. The resources
required to combine multi-spectral fluorescent reagents,
immunochemicals, and other assay components into a
kit that retains the sensitivity and specificity of each component
creates a large overhead in HCS assay design. Therefore,
it is beneficial to access HCS reagent kits comprised of
validated combinations of reagents, removing the burden of
assay design from the end-user.
INFORMATICS/BIOINFORMATICS
A key component of the integrated and distributed capability
of advanced HCS is the availability of a toolbox that
accesses most ofthe HCS platform right on the desktop ofthe
scientists who need this capability the most. This software
package includes Bio Application software, which allows
archiving, retrieval, and visualization of image and associated
numerical assay output features, an application that
permits reprocessing of acquired image data, software for
rapid assay design and analyses of considerably large data
sets, and a program that allows rapid discovery of ranked
relationships from the literature to qualify targets, genes,
proteins, cells, diseases, compounds, and screening results
(e.g., CellSpaceKnowledge Miner). Many investigators can
therefore run ''in silico'' experiments in parallel that reprocess
existing data sets with new parameters, mine data, design
new assays, and verify information and knowledge from
available literature, all while the HCS instrumentation is
running new screens. Thus, HCS can scale beyond the
number of physical instruments.
An optimal HCS solution must have an open architecture
design that ensures image data obtained from HCS instruments,
as well as other imaging microscope systems, can be
archived, managed, analyzed, and explored on a single
platform connected to multiple personal workstations. This
is an important feature, since many HCS assays grow out of
small-scale biology experiments using research-imaging
systems including laser scanning confocal microscopes.
In addition, the open architecture allows the incorporation
of powerful software tools available from other vendors
such as Spotfire (Somerville, MA) DecisionSite,IDBS
(Guildford, UK) Activity Base, and MDL ISIS (MDL Information
Systems, San Leandro, CA) or contextual linking
of HCS data to genomic and proteomic data using federated
databases.
The general trend of increased use of automated methods
for generating data, as well as the large amount of rich
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