Reduction of autofluorescence on DNA microarrays and slide surfaces by treatment with sodium borohydride.
Microarray technology is currently being used extensively in functional genomics research and modern drug discovery and development. Henceforward, tremendous application potential for this technology exists in the fields of clinical diagnostics and prognostics, pathology, and toxicology for high-throughput analysis of "disease" gene expression. However, the major hurdle now in this technology is not the performance of the arrays but rather the efficient reproducibility of the hybridization signal intensity in a fluorescence-based analysis. The sensitivity of fluorescence detection on an array is to a large extent limited by the amount of background signal arising due to nonspecifically bound probes and fluorescence that is intrinsically associated with the chip substrate and/or the attached target DNA, the so-called autofluorescence. Here, we describe a simple and efficient method to reduce autofluorescence from undetermined sources on coated glass slides with and without DNA arrays. This sodium borohydride-mediated reduction process resulted in significantly lower and more even background fluorescence. This in turn extended the dynamic range of detection and reduced the average coefficient of variation of fluorescent signal ratios on DNA microarrays in addition to improving the detection of genes that are expressed at a low level.
Raghavachari
Nalini
N
Biochemical Technologies, Corning Inc., NY 14831, USA. raghavacn@corning.com
Bao
Yijia P
YP
Li
Guangshan
G
Xie
Xinying
X
Müller
Uwe R
UR
D001894:Borohydrides / Q000737:chemistry*; D001921:Brain / Q000378:metabolism; D018076:DNA, Complementary / Q000032:analysis* / Q000235:genetics; D005453:Fluorescence; D020869:Gene Expression Profiling / Q000295:instrumentation / Q000379:methods*; D005898:Glass; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D001894:Borohydrides; D018076:DNA, Complementary; C025364:sodium borohydride