Details
| Original language | English |
|---|---|
| Pages (from-to) | 556-572 |
| Number of pages | 17 |
| Journal | Biology |
| Volume | 4 |
| Issue number | 3 |
| Publication status | Published - 11 Sept 2015 |
Abstract
DNA-Microarrays have become a potent technology for high-throughput analysis of genetic regulation. However, the wide dynamic range of signal intensities of fluorophore-based microarrays exceeds the dynamic range of a single array scan by far, thus limiting the key benefit of microarray technology: parallelization. The implementation of multi-scan techniques represents a promising approach to overcome these limitations. These techniques are, in turn, limited by the fluorophores’ susceptibility to photobleaching when exposed to the scanner’s laser light. In this paper the photobleaching characteristics of cyanine-3 and cyanine-5 as part of solid state DNA microarrays are studied. The effects of initial fluorophore intensity as well as laser scanner dependent variables such as the photomultiplier tube’s voltage on bleaching and imaging are investigated. The resulting data is used to develop a model capable of simulating the expected degree of signal intensity reduction caused by photobleaching for each fluorophore individually, allowing for the removal of photobleaching-induced, systematic bias in multi-scan procedures. Single-scan applications also benefit as they rely on pre-scans to determine the optimal scanner settings. These findings constitute a step towards standardization of microarray experiments and analysis and may help to increase the lab-to-lab comparability of microarray experiment results.
Keywords
- Bioanalytics, Bioinformatics, Cyanine dye, DNA, Fluorophore, Microarray, Photobleaching
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- General Biochemistry,Genetics and Molecular Biology
- Immunology and Microbiology(all)
- General Immunology and Microbiology
- Agricultural and Biological Sciences(all)
- General Agricultural and Biological Sciences
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In: Biology, Vol. 4, No. 3, 11.09.2015, p. 556-572.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - The Impact of Photobleaching on Microarray Analysis
AU - von der Haar, Marcel
AU - Preuß, John Alexander
AU - von der Haar, Kathrin
AU - Lindner, Patrick
AU - Scheper, Thomas
AU - Stahl, Frank
PY - 2015/9/11
Y1 - 2015/9/11
N2 - DNA-Microarrays have become a potent technology for high-throughput analysis of genetic regulation. However, the wide dynamic range of signal intensities of fluorophore-based microarrays exceeds the dynamic range of a single array scan by far, thus limiting the key benefit of microarray technology: parallelization. The implementation of multi-scan techniques represents a promising approach to overcome these limitations. These techniques are, in turn, limited by the fluorophores’ susceptibility to photobleaching when exposed to the scanner’s laser light. In this paper the photobleaching characteristics of cyanine-3 and cyanine-5 as part of solid state DNA microarrays are studied. The effects of initial fluorophore intensity as well as laser scanner dependent variables such as the photomultiplier tube’s voltage on bleaching and imaging are investigated. The resulting data is used to develop a model capable of simulating the expected degree of signal intensity reduction caused by photobleaching for each fluorophore individually, allowing for the removal of photobleaching-induced, systematic bias in multi-scan procedures. Single-scan applications also benefit as they rely on pre-scans to determine the optimal scanner settings. These findings constitute a step towards standardization of microarray experiments and analysis and may help to increase the lab-to-lab comparability of microarray experiment results.
AB - DNA-Microarrays have become a potent technology for high-throughput analysis of genetic regulation. However, the wide dynamic range of signal intensities of fluorophore-based microarrays exceeds the dynamic range of a single array scan by far, thus limiting the key benefit of microarray technology: parallelization. The implementation of multi-scan techniques represents a promising approach to overcome these limitations. These techniques are, in turn, limited by the fluorophores’ susceptibility to photobleaching when exposed to the scanner’s laser light. In this paper the photobleaching characteristics of cyanine-3 and cyanine-5 as part of solid state DNA microarrays are studied. The effects of initial fluorophore intensity as well as laser scanner dependent variables such as the photomultiplier tube’s voltage on bleaching and imaging are investigated. The resulting data is used to develop a model capable of simulating the expected degree of signal intensity reduction caused by photobleaching for each fluorophore individually, allowing for the removal of photobleaching-induced, systematic bias in multi-scan procedures. Single-scan applications also benefit as they rely on pre-scans to determine the optimal scanner settings. These findings constitute a step towards standardization of microarray experiments and analysis and may help to increase the lab-to-lab comparability of microarray experiment results.
KW - Bioanalytics
KW - Bioinformatics
KW - Cyanine dye
KW - DNA
KW - Fluorophore
KW - Microarray
KW - Photobleaching
UR - http://www.scopus.com/inward/record.url?scp=84941358397&partnerID=8YFLogxK
U2 - 10.3390/biology4030556
DO - 10.3390/biology4030556
M3 - Article
C2 - 26378589
AN - SCOPUS:84941358397
VL - 4
SP - 556
EP - 572
JO - Biology
JF - Biology
IS - 3
ER -