Human T lymphocytes were stimulated using phorbol myristate acetate and ionomycin. Twenty-four hours post-activation the cells were harvested for DNA content and for measurements using a newly developed cell profiling system employing dielectrophoresis. This system provides individual cell size and dielectrophoresis data for statistically relevant numbers of control and activated cells. From this it was determined that the mean membrane specific capacitance decreased from 13.49 (+/- 4.72) mF/m(2) to 10.62 (+/- 5.13) mF/m(2). This can be related to a 21.3% reduction in the effective membrane surface area associated with membrane topography (e.g. reduction of membrane associated microvilli, blebs and folding), or to other changes of membrane architecture, following cell activation. From cytometric determinations of DNA content, it was concluded that these effects were related to a 3.0-fold decrease of cells in S-phase, and a 1.5-fold increase in G1 cells. This work demonstrates the powerful potential of using dielectrophoresis as a noninvasive tool to follow physiological changes that accompany transmembrane signaling events.

Global gene expression analysis using Affymetrix GeneChip Human Transcriptome Array (HTA) 2.0 combined with Affymetrix Transcriptome Analysis Console (TAC) Software empowers scientists not only to gain information about RNA expression at the gene‐level, but also about RNA alternative splicing at the exon‐level. Real‐time PCR is routinely used to verify microarray results due to its high sensitivity and wide dynamic range, and was used in this study to validate Affymetrix GeneChip HTA 2.0 data. Gene‐level expression data for the classic MAQC A and MAQC B samples (used by the MicroArray Quality Control Consortium) were validated with commercially available primers and design software. Excellent fold‐change correlation of microarray and qPCR data were observed using USB VeriQuest Probe real‐time PCR (R=0.96, slope=0.70). For alternative splicing validation of Affymetrix GeneChip HTA 2.0, total RNA from three different tissues (liver, muscle and brain) were used. Exon level information provided by TAC Software was utilized to design custom PCR primers for USB VeriQuest SYBR Green real‐time PCR. All alternative splicing events selected for validation were confirmed by real‐time PCR. We show excellent inter‐platform concordance between Affymetrix microarray and VeriQuest real‐time PCR data. Grant Funding Source: Supported by Affymetrix, Inc.

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