Genome-wide platelet RNA profiling in clinical samples.
Human blood platelets are anucleate cells that contain minute amounts of translational active mRNA. Investigation of the gene expression profile by microarray analysis has become an excellent tool for better understanding of normal and pathological platelet function. Its use, however, is often limited by the low yield of megakaryocytic-derived mRNA, the possible contamination with leukocytes during platelet preparation and the small platelet volume in clinical settings, especially in pediatric patients. In this chapter, we present a protocol for the isolation of leukocyte-depleted platelet samples in clinical settings and an optimized procedure for transcript profiling, using the Agilent oligo microarray technology. In particular, we discuss the special aspects of platelet purification when working with blood sample volumes not exceeding 3-5 ml, which is typical in pediatric patients and we furthermore provide detailed information for transcript profiling of extremely small amounts of platelet RNA.
Institute of Transfusion Medicine and Immunology, University of Heidelberg, Medical Faculty of Mannheim, Mannheim, Germany.
Angelika
A
Schedel
Nina
N
Rolf
D001792:Blood Platelets / Q000378:metabolism*; D001800:Blood Specimen Collection / Q000379:methods; D020869:Gene Expression Profiling / Q000379:methods*; D015894:Genome, Human; D006801:Humans; D008533:Megakaryocytes / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012333:RNA, Messenger / Q000096:biosynthesis*
D012333:RNA, Messenger