Direct measurement of multiple mRNAs in nerve growth factor-induced PC12 cells using electrophoretic tags to monitor biomarkers of neuronal differentiation in 96-well format.

Pheochromocytoma-12 (PC12) cells recapitulate the program of neuronal differentiation by developing neurites after about 12 days of nerve growth factor (NGF) treatment. This model can be used to evaluate the neuroprotective/neurotrophic effect of compounds. Specific mRNAs such as cfos and c-jun are early biomarkers of the irreversible commitment into the differentiation program as they appear after only 30–40 min of NGF treatment. Monitoring the level of these mRNAs instead of the neurite outgrowth dramatically reduces the time needed to identify the drug potential of compounds. The electrophoretic tags, or eTag™ reporters (ACLARA Biosciences, Inc., Mountain View, CA), are a new class of fluorescent reporters that have unique migration properties in capillary electrophoresis, which allows for their separation and identification. (The eTag Multiplex Invader® Assay and products incorporate Invader® technology and Cleavase® enzyme licensed for use from Third Wave Technologies, Inc. [Madison, WI] for multiple...