title: Real-time reverse transcription-PCR quantitation of substance P receptor (NK-1R) mRNA.
abstract: The substance P (SP)-preferring receptor, neurokinin-1 receptor (NK-1R), has an important role in inflammation, immune regulation, and viral infection. We applied a newly developed real-time reverse transcription (RT)-PCR assay to quantify NK-1R mRNA in human neuronal cell line (NT-2N), a human B-cell line (IM9), monocyte-derived macrophages (MDM), peripheral blood lymphocytes (PBL), and human astroglioma cells (U87 MG). The NK-1R real-time RT-PCR assay has a sensitivity of 100 mRNA copies, with a dynamic range of detection between 10(2) and 10(7) copies of NK-1R gene transcripts per reaction. This assay is highly reproducible, with an intraassay coefficient variation of threshold cycle (Ct) of less than 1.9%. The NK-1R real-time RT-PCR is highly sensitive for quantitative determination of NK-1R mRNA in human immune cells (MDM and PBL) that express low levels of NK-1R mRNA. In addition, the assay has the ability to accurately quantitate the dynamic changes in NK-1R mRNA expression in interleukin-1beta-stimulated U87 MG. These data indicate that the NK-1R real-time RT-PCR has potential for a wide application in investigation of NK-1R expression at the mRNA level under physiological and pathological conditions in both the central nervous system and the immune system.
authors.affiliation: Division of Allergy and Immunology, Joseph Stokes Jr. Research Institute at the Children's Hospital of Philadelphia, Department of Pediatrics, University of Pennsylvania Medical School, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Jian-Ping
authors.initials: JP
authors.lastname: Lai
authors.forename: Steven D
authors.initials: SD
authors.lastname: Douglas
authors.forename: Yan-Jian
authors.initials: YJ
authors.lastname: Wang
authors.forename: Wen-Zhe
authors.initials: WZ
authors.lastname: Ho
mesh_terms: D002478:Cells, Cultured; D005786:Gene Expression Regulation; D006801:Humans; D016014:Linear Models; D012333:RNA, Messenger / Q000032:analysis; D018040:Receptors, Neurokinin-1 / Q000032:analysis* / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
chemical_list: D012333:RNA, Messenger; D018040:Receptors, Neurokinin-1