High-sensitivity detection of DNA hybridization on microarrays using resonance light scattering.
The application of resonance light scattering (RLS) particles for high-sensitivity detection of DNA hybridization on cDNA microarrays is demonstrated. Arrays composed of approximately 2000 human genes ("targets") were hybridized with colabeled (Cy3 and biotin) human lung cDNA probes at concentrations ranging from 8.3 ng/microL to 16.7 pg/microL. After hybridization, the arrays were imaged using a fluorescence scanner. The arrays were then treated with 80-nm-diameter gold RLS Particles coated with anti-biotin antibodies and imaged in a white light, CCD-based imaging system. At low probe concentrations, significantly more genes were detected by RLS compared to labeling by Cy3. For example, for hybridizations with a probe concentration of 83.3 pg/microL, approximately 1150 positive genes were detected using RLS compared to approximately 110 positive genes detected with Cy3. In a differential gene expression experiment using human lung and leukemia RNA samples, similar differential expression profiles were obtained for labeling by RLS and fluorescence technologies. The use of RLS Particles is particularly attractive for detection and identification of low-abundance mRNAs and for those applications in which the amount of sample is limited.
Bao
Paul
P
Biochemical Technologies, Science and Technology Division, Coming Incorporated, Coming, New York 14831, USA.
Frutos
Anthony G
AG
Greef
Charles
C
Lahiri
Joydeep
J
Muller
Uwe
U
Peterson
Todd C
TC
Warden
Laurence
L
Xie
Xinying
X
D015342:DNA Probes / Q000737:chemistry; D020869:Gene Expression Profiling / Q000379:methods; D006801:Humans; D008027:Light; D008168:Lung / Q000737:chemistry; D009693:Nucleic Acid Hybridization / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012334:RNA, Neoplasm / Q000032:analysis; D012542:Scattering, Radiation; D012680:Sensitivity and Specificity
D015342:DNA Probes; D012334:RNA, Neoplasm