Gold nanoparticle probe-based gene expression analysis with unamplified total human RNA.
Microarray-based gene expression analysis plays a pivotal role in modern biology and is poised to enter the field of molecular diagnostics. Current microarray-based gene expression systems typically require enzymatic conversion of mRNA into labeled cDNA or cRNA. Conversion to cRNA involves a target amplification step that overcomes the low sensitivity associated with commonly used fluorescent detection methods. Herein, we present a novel enzyme-free, microarray-based gene expression system that uses unamplified total human RNA sample as the target nucleic acid. The detection of microarray-bound RNA molecules is accomplished by targeting the poly-A tail with an oligo-dT20 modified gold nanoparticle probe, signal amplification by autometallography, and subsequent measurement of nanoparticle-mediated light scattering. The high sensitivity afforded by the nanoparticle probes allows differential gene expression from as little as 0.5 microg unamplified total human RNA in a 2 h hybridization without the need for elaborate sample labeling steps.
Huber
Martin
M
Nanosphere Inc., 4088 Commercial Ave., Northbrook, IL 60062, USA.
Wei
Tai-Fen
TF
Müller
Uwe R
UR
Lefebvre
Phil A
PA
Marla
Sudhakar S
SS
Bao
Y Paul
YP
D015374:Biosensing Techniques; D020869:Gene Expression Profiling / Q000379:methods*; D006046:Gold / Q000737:chemistry*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D015345:Oligonucleotide Probes / Q000737:chemistry*; D012333:RNA, Messenger / Q000032:analysis* / Q000378:metabolism; D015203:Reproducibility of Results
D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D006046:Gold