Protocol

Cell-Based Microarrays

Volume 706 of the series Methods in Molecular Biology pp 191-206

Date:

Microfluidic Image Cytometry

  • Ken-ichiro KameiAffiliated withDepartment of Molecular & Medical Pharmacology, David Geffen School of Medicine, University of California Email author 
  • , Jing SunAffiliated withDepartment of Molecular & Medical Pharmacology, David Geffen School of Medicine, University of California
  • , Hsian-Rong TsengAffiliated withDepartment of Molecular & Medical Pharmacology, David Geffen School of Medicine, University of California
  • , Robert DamoiseauxAffiliated withMolecular Screening Shared Resource, David Geffen School of Medicine, University of California

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Abstract

Cell-based arrays offer powerful tools for genomics/proteomics and drug discovery, and are widely applicable for most cell lines. However, it is challenging to apply cell-based arrays for in vitro diagnosis due to limited amount of patient samples. Here, we utilized and demonstrated microfluidic image cytometry (MIC), capable of quantitative, single-cell profiling of multiple signaling molecules using only 300–3,000 cells from clinical brain tumor specimens for in vitro molecular diagnosis. First, we characterized the PI3K/AKT/mTOR pathway, which is often over-activated in the brain tumors, in U87 brain tumor cell lines by measuring EGFR, PTEN, pAKT, and pS6 with a MIC platform, and applied this measurement to clinical brain tumor specimens. In conjunction with statistical analysis, we were able to characterize extensive inter- and intra-tumoral molecular heterogeneity.

Key words

Microfluidic image cytometry molecular diagnosis glioblastoma Pi3k signaling pathway immunofluorescence biostatistics