title: Expanding applications of protein analysis using proximity ligation and qPCR.
abstract: The correlation of gene and protein expression changes in biological systems has been hampered by the need for separate sample handling and analysis platforms for nucleic acids and proteins. In contrast to the simple, rapid, and flexible workflow of quantitative PCR (qPCR) methods, which enable characterization of several classes of nucleic acid biomarkers (i.e. DNA, mRNA, and microRNAs), protein analysis methods such as Western blotting are cumbersome, laborious, and much less quantitative. However, TaqMan(R) Protein Assays, which use the proximity ligation assay (PLA) technology, now expand the range of qPCR applications to include the direct detection of proteins through the amplification of a surrogate DNA template after antibody binding. Here we describe an integrated qPCR approach for measuring relative changes in gene and protein expression from the same starting sample and on a single analytical platform that pairs TaqMan Gene Expression (GEx) Assays with TaqMan Protein Assays. We have monitored the changes in mRNA, microRNA, and protein expression of relevant biomarkers in the pluripotent human embryonal carcinoma cell line, NTERA2, upon differentiation to neuronal cells. In addition, TaqMan Protein Assays have been used to monitor protein expression in induced pluripotent stem cells (iPSC) that have been reprogrammed from human somatic cells. The data presented establishes a general paradigm utilizing real-time PCR instruments and reagents for studying the relationship between the stem cell transcriptome and proteome.
authors.affiliation: Applied Biosystems, Part of Life Technologies, 850 Lincoln Centre Drive, Foster City, CA 94404, USA. Elana.Swartzman@lifetech.com
authors.forename: Elana
authors.initials: E
authors.lastname: Swartzman
authors.forename: Mark
authors.initials: M
authors.lastname: Shannon
authors.forename: Pauline
authors.initials: P
authors.lastname: Lieu
authors.forename: Shiaw-Min
authors.initials: SM
authors.lastname: Chen
authors.forename: Chad
authors.initials: C
authors.lastname: Mooney
authors.forename: Eric
authors.initials: E
authors.lastname: Wei
authors.forename: Julie
authors.initials: J
authors.lastname: Kuykendall
authors.forename: Rouying
authors.initials: R
authors.lastname: Tan
authors.forename: Tina
authors.initials: T
authors.lastname: Settineri
authors.forename: Levente
authors.initials: L
authors.lastname: Egry
authors.forename: David
authors.initials: D
authors.lastname: Ruff
mesh_terms: D002454:Cell Differentiation / Q000187:drug effects; D045744:Cell Line, Tumor; D019411:Clinical Laboratory Techniques; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000378:metabolism; D035683:MicroRNAs / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D014212:Tretinoin / Q000494:pharmacology
chemical_list: D035683:MicroRNAs; D012333:RNA, Messenger; D014212:Tretinoin