After you have identified genes with altered expression in your experimental group, the expression
levels of a sub-set of these should be validated. Most journals will not publish without some validation
of the gene sets identified in microarray studies.
Real-time PCR (two systems available on site)
Quantitative RT-PCR (multiplex with internal standard best option for RT-PCR)
NB: Probe match on NetAffx (Affymetrix website) can be used to check your oligonucleotide against
the target sequence. NetAffx also contains all sequence information for oligonucleotides used on the
GeneChip™ arrays. These can be used in your validation studies as the oligonucleotides have been
selected for optimal, specific hybridization to the target sequence. The signals for each oligonucleotide
and its paired mismatch are in the CHP files associated with each sample. This allows you to check the
efficiency and specificity of hybridization for each oligonucleotide to each of the samples that you have
You may wish to extend your studies to examine protein expression levels of the genes identified as
‘interesting’ in your microarray study (e.g. Westerns, IHC, etc).