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Series GSE18161 Query DataSets for GSE18161
Status Public on Sep 22, 2009
Title Washing scaling of microarray expression
Organism Homo sapiens
Experiment type Expression profiling by array
Summary Post-hybridization washing is an essential part of microarray experiments. Both, the quality of the experimental washing protocol and the adequate consideration of washing in intensity calibration ultimately affect the quality of the expression estimates extracted from the microarray intensities. We conducted experiments on GeneChip microarrays with altered protocols for washing, scanning and staining to study the probe-level intensity changes as a function of washing cycles. Particularly, three Affymetrix GeneChip HGU133plus2 arrays were hybridized and equilibrated for 16 hours in the hybridization oven. For one of the three arrays washing and staining was performed according to the manufacturer’s instructions. For another array the first scan was done immediately after low stringent wash and staining without intermitting stringent washing. Then, the array was stringently washed and scanned in alternating order three more times where each washing step consists of a definite number of washing cycles. The third array was low stringently washed followed by two stringent washing cycles and staining before the first scan. Subsequently it was analogously processed as array A. All three chips are repeatedly processed in a second series of alternating wash/scan-cycles which was performed using the same protocol for each chip as in the first series as described above. As in the first series the arrays were also stained a second time to compensate for any loss of bleached fluorescent dye. Analysis of the washing kinetics shows that the signal-to-noise ratio doubles roughly every ten stringent washing cycles. Washing can be characterized by time-dependent rate constants which reflect the heterogeneous character of target binding to microarray probes. We propose an empirical washing function which estimates the survival of probe bound targets. The washing function allows calibrating probe intensities for the effect of washing. On a relative scale, proper calibration for washing markedly increases expression measures especially in the limit of small and large values.
 
Overall design The aim of the study was to understand the probe-level intensity changes as a function of washing cycles. For this experiment RNA was prepared from a cancer cell line and used for probe synthesis. Hybridisation to three Affymetrix microarrays was identically performed with aliquotes of one hybridization mix from a single RNA sample. The protocols for washing, scanning and staining were altered.
 
Contributor(s) Binder H, Krohn K, Burden C
Citation(s) 20509934
Submission date Sep 18, 2009
Last update date Mar 25, 2019
Contact name Knut Krohn
E-mail(s) knut.krohn@medizin.uni-leipzig.de
Organization name University Leipzig
Department Medizinische Fakultät
Lab CU DNA im SIKT
Street address Philipp-Rosenthal-Straße 55
City Leipzig
ZIP/Postal code 04103
Country Germany
 
Platforms (1)
GPL570 [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array
Samples (18)
GSM453956 A1, 0x st_wash, 1st series
GSM453957 A2, 2x st_wash, 1st series
GSM453958 A3, 7x st_wash, 1st series
Relations
BioProject PRJNA119465

Washing protocol header descriptions
title
CHP file
CEL file
stringent wash cycles (total)
initial stringent wash cycles before first staining (first series)
staining (first series)
additional stringent wash cycles after first staining (first series)
initial stringent wash cycles before second staining (second series)
staining (second series)
additional stringent wash cycles after second staining (second series)
-

Data table
title CHP file CEL file stringent wash cycles (total) initial stringent wash cycles before first staining (first series) staining (first series) additional stringent wash cycles after first staining (first series) initial stringent wash cycles before second staining (second series) staining (second series) additional stringent wash cycles after second staining (second series) -
A1, 0x st_wash, 1st series A1.mas5.CHP A1.CEL 0x 0x 3 rounds scanning
A2, 2x st_wash, 1st series A2.mas5.CHP A2.CEL 2x 0x 3 rounds 2x scanning
A3, 7x st_wash, 1st series A3.mas5.CHP A3.CEL 7x 0x 3 rounds 7x scanning
A4, 17x st_wash, 1st series A4.mas5.CHP A4.CEL 17x 0x 3 rounds 17x scanning
A5, 17x st_wash, 2nd series A5.mas5.CHP A5.CEL 17x 0x 3 rounds 17x 0x 3 rounds scanning
A6, 19x st_wash, 2nd series A6.mas5.CHP A6.CEL 19x 0x 3 rounds 17x 0x 3 rounds 2x scanning
A7, 24x st_wash, 2nd series A7.mas5.CHP A7.CEL 24x 0x 3 rounds 17x 0x 3 rounds 7x scanning
A8, 34x st_wash, 2nd series A8.mas5.CHP A8.CEL 34x 0x 3 rounds 17x 0x 3 rounds 17x scanning
B1, 2x st_wash, 1st series B1.mas5.CHP B1.CEL 2x 2x 3 rounds scanning
B2, 4x st_wash, 1st series B2.mas5.CHP B2.CEL 4x 2x 3 rounds 2x scanning
B3, 9x st_wash, 1st series B3.mas5.CHP B3.CEL 9x 2x 3 rounds 7x scanning
B4, 19x st_wash, 1st series B4.mas5.CHP B4.CEL 19x 2x 3 rounds 17x scanning
B5, 21x st_wash, 2nd series B5.mas5.CHP B5.CEL 21x 2x 3 rounds 17x 2x 3 rounds scanning
B6 23x st_wash, 2nd series B6.mas5.CHP B6.CEL 23x 2x 3 rounds 17x 2x 3 rounds 2x scanning
B7, 28x st_wash, 2nd series B7.mas5.CHP B7.CEL 28x 2x 3 rounds 17x 2x 3 rounds 7x scanning
B8, 38x st_wash, 2nd series B8.mas5.CHP B8.CEL 38x 2x 3 rounds 17x 2x 3 rounds 17x scanning
C1, 6x st_wash, 1st series C1.mas5.CHP C1.CEL 6x 6x 3 rounds scanning
C2, 12x st_wash, 2nd series C2.mas5.CHP C2.CEL 12x 6x 3 rounds 0x 6x 3 rounds scanning

Total number of rows: 18


Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE18161_RAW.tar 119.4 Mb (http)(custom) TAR (of CEL, CHP)
Processed data included within Sample table
Processed data provided as supplementary file
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