title: Isolation of circulating microRNA associated with RNA-binding protein.
abstract: Circulating microRNAs (miRNAs) have been recently detected in extracellular body fluids and proved themselves as promising biomarkers for a broad spectrum of diseases. The techniques to isolate, detect, and characterize extracellular miRNAs vary significantly from report to report. In this chapter we describe a consistent, efficient and highly reproducible protocol for isolation and detection of microvesicles-free circulating miRNA from human blood plasma or cell culture media. Furthermore, since exosomes-free circulating miRNAs are associated with Argonaut proteins (the same proteins to which miRNAs are bound inside the cell), we provide a protocol for immunoprecipitation of AGO2 associated miRNAs.
authors.affiliation: Molecular Epidemiology, German Cancer Research Center, Heidelberg, Germany.
authors.forename: Andrey
authors.initials: A
authors.lastname: Turchinovich
authors.forename: Ludmila
authors.initials: L
authors.lastname: Weiz
authors.forename: Barbara
authors.initials: B
authors.lastname: Burwinkel
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry; D060565:Argonaute Proteins / Q000302:isolation & purification* / Q000378:metabolism; D017077:Culture Media, Conditioned / Q000737:chemistry; D006801:Humans; D047468:Immunoprecipitation; D007963:Leukocytes, Mononuclear / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D035683:MicroRNAs / Q000145:classification / Q000302:isolation & purification*; D020411:Oligonucleotide Array Sequence Analysis; D011485:Protein Binding; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: C466422:AGO2 protein, human; D000911:Antibodies, Monoclonal; D060565:Argonaute Proteins; D017077:Culture Media, Conditioned; D035683:MicroRNAs