Snapshots Live Cytokines & Growth Factors June 11, 2003

Agenda Introduction Cell- and Protein-Based Assays Microarrays Conclusions

Cytokines & Growth Factors Diverse roles in development and disease Study includes receptors as well as cytokine and growth factor ligands Many different molecule families and family members Introduction

Most Frequently Studied Molecules Introduction

Most Popular Assays Introduction Cell-Based Assays Protein-Based Assays RNA-Based Assays Growth inhibition or proliferation ELISAReal-time PCR Flow cytometryWestern blottingEnd-point PCR Fluorescent microscopy Immuno- precipitation DNA microarrays

Cell- and Protein-Based Assays Key Requirements Signal-to-noise Detection equipment Sensitivity High-throughput Critical Issues All product categories – species selection Antibodies – use with fixed tissues Immunoassay kits – shelf life Recombinant proteins – molecule selection

Microarray Formats Used in RNA-Based Assays Microarrays

Sources of Microarrays for Cytokine Research Microarrays TaKaRa Biomedical 1% Radius Biosciences 1% OriGene Technologies 1% Mergen 1% Genemed Biotechnologies 1% Sigma-Genosys 2% Perkin Elmer Life Sciences 2% Genomic Solutions 2% MWG Biotech 3% Amersham Biosciences 6% Agilent Technologies 6% BD Biosciences Clontech 8% SuperArray 9% Other 20% Affymetrix 37%

Conclusions Cytokine researchers study a wide diversity of molecules and species; off-the-shelf assay kits and reagents are often hard to find. Growth inhibition/proliferation, ELISA and real-time PCR are the most popular assays for cytokine and growth factor research. Microarray users identified difficulty with data interpretation, specificity and sensitivity as the most common problems encountered.