Microarray analysis using a limited amount of cells.
cDNA microarray technology is widely used in various biological and medical disciplines to determine gene expression profiles. Unfortunately, this technology requires a large quantity of input RNA. Since there is an increasing need for more precise analyses of defined cell subpopulations with low cell counts, working protocols using a minimal number of input cells are required. Optimal RNA isolation and its accurate amplification are crucial to the success of these protocols. The HL-60 cell line was used in the search for a suitable protocol that can be used for clinical samples of CD34+ haematopoietic cells obtained from bone marrow. The goal was to discover the best method for isolating and amplifying RNA from a small number of cells. Our evaluation of various methods and kits available in the market revealed that the combination of RNAqueous Kit for RNA isolation and the SenseAmp Plus Kit for one-round RNA amplification produced the best results. This article presents a verified protocol describing a reliable and reproducible method for obtaining enough input RNA for microarray experiments when the number of cells is limited.
Peterková
M
M
Department of Internal Hematooncology, Faculty of Informatics, Centre for Biomedical Image Analysis, Masaryk University, Brno, Czech Republic. peterkov@fi.muni.cz
Koutná
I
I
Tesarová
L
L
Potesilová
M
M
Kozubek
M
M
Hrabcáková
V
V
Klabusay
M
M
Doubek
M
M
Mayer
J
J
D002478:Cells, Cultured; D018922:HL-60 Cells; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification
D012313:RNA