By Nobuyoshi Kosaka
microRNAs (miRNAs) are small non-coding RNAs that keep an eye on numerous organic phenomena, similar to improvement and homeostasis. The dysregulation of miRNA ends up in ailment development, fairly of melanoma. In Circulating MicroRNAs: tools and Protocols, specialist researchers within the box aspect fresh advances within the isolation, purification and research of circulating miRNAs from quite a few assets for study. The e-book is split into 3 major subject matters. the 1st part consists of the research of secretory miRNAs in cell-cell conversation, and the second one, the examine of circulating miRNAs in physique fluids. The final describes the radical strategies used to review circulating miRNAs. Written within the hugely profitable Methods in Molecular Biology™ sequence layout, chapters comprise introductions to their respective themes, lists of the mandatory fabrics and reagents, step by step, effectively reproducible laboratory protocols, and key tips about troubleshooting and warding off recognized pitfalls.
Authoritative and practical, Circulating MicroRNAs: tools and Protocols seeks to help scientists in facing the hot advances of RNAi know-how from the bench to the bedside.
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Extra resources for Circulating MicroRNAs: Methods and Protocols
Exosomes also contain a specific repertoire of microRNAs and mRNAs that differs from the miRNA and mRNA repertoire of the cell of origin [4, 5]. Exosomes can be used by immune cells and other cell types as devices for communication via the direct exchange of RNA and proteins between cells separated at a distance . Recently, we demonstrated that the transfer of exosomes from a T cell to an APC is promoted by the formation of a functional synapse, therefore facilitating the exchange of exosomal proteins and microRNAs .
22 μm filters. 3. 45 μm, antibodies for blotting exosomal markers: antiCD81, anti-CD63. 4. BCA protein quantification: BCA protein assay kit, 2 mg/ml BSA, 96-well flat-bottom plates, spectrophotometer. 5. FACS analysis of exosomes: Ultra-clean PBS, PE mouse antihuman CD63 (BD Biosciences), BD FACSCanto Flow Cytometer (Becton Dickinson), Count Bright Absolute counting beads (Invitrogen). 6. Cell cocultures: PBS, blue fluorescent cell tracker CMAC, superantigen endotoxin E, 96-well round-bottom plates.
References 1. Wang K, Zhang S, Weber J et al (2010) Export of microRNAs and microRNA-protective protein by mammalian cells. Nucleic Acids Res 38:7248–7259 2. Arroyo JD, Chevillet JR, Kroh EM et al (2011) Argonaute2 complexes carry a population of circulating microRNAs independent of vesicles in human plasma. Proc Natl Acad Sci USA 108:5003–5008 3. Valadi H, Ekstrom K, Bossios A et al (2007) Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells. Nat Cell Biol 9:654–659 4.