HTS FLOW CYTOMETRY, SMALL-MOLECULE DISCOVERY, AND THE NIH MOLECULAR LIBRARIES INITIATIVE - Flow Cytometry in Drug Discovery and Development - page 75

5-10 samples perminute. TheHyperCyt platform for HTSflowcytometry, currently
available at 30 or more sites around the world through IntelliCyt [33], can acquire
samples at rates up to 40 wells per minute. These sampling rates are currently
compatible with throughput from several thousands to many tens of thousands of
samples per day, depending upon the number of hours and extent of automation.
Sites actively reporting usage of flow cytometry in a discovery mode for primary
screens in addition to UNMCMD include (1) Vivia [34], a successor to Novasite
that performs reprofiling of approved drugs; (2) Rigel that screens SiRNA [35]; (3)
and the Center for Chemical Genomics at the University of Michigan Life Science
Institute. To our knowledge, the Rigel site is unique in having a fully automated and
integrated platform that was developed in collaboration with IntelliCyt and Agilent.
A partnership between IntelliCyt and Beckman Coulter [36], the international
distributor of the HyperCyt platform, links the Beckman sample handling Biomek
platform, HyperCyt, and the CyAn flow cytometer that was recently acquired by
Beckman Coulter.
Essential opportunities of flow cytometry for small-molecule discovery are
summarized in Table 4.2. Flow cytometry is well known as a multicolor technology
that can be used in multiplexing or mutiparameter analysis (the analogue to high-
content analysis in microscopy), as well as the homogeneous discrimination of ligand
binding and macromolecular assemblies.
The versatility of the flow cytometry platform is remarkable and well known.
Capabilities range from 10 color immunological assays and extend to most cell-

based assays that can be performed fluorimetrically or microscopically, including cell
proliferation, differentiation, death, signaling, adhesion, ligand binding, transport,
and others. There are published protocols, stains, and markers for surface, cytoplas-
mic, and nuclear components. Flow cytometry excels at phenotypic assays, such as
antibody staining and light scatter (size and shape) discrimination used on complex

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TABLE 4.2 Flow Cytometry Features Useful for HTS

Feature Utility Application

Homogeneous detection Real-time discrimination

of particle bound from
unbound molecules

Ligand binding or molecular
assemblies. Conserves
precious reagents on beads

Multiparameter 10 or more colors and scatter

signals

High content, e.g., phospho-
protein analysis

Multiplexing and
barcoding

Two or more colors and scatter
signals used to distinguish
multiple particle targets

Suspension arrays for multiple
assemblies or multiple cell
targets

Acquisition rate/digital
signal processing

50,000 events per second Thousands of flow cytometric

assays accessible to high
throughput

Size and shape
discrimination

Particle sizes ranging from
submicron to tens of
micrometers; changes
in shape/morphology

Cells, particles, solubility
screening of compound
libraries