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Sample GSM115773 Query DataSets for GSM115773
Status Public on Jul 14, 2006
Title Prostate_03-115E_GL4
Sample type RNA
 
Channel 1
Source name Prostate_normal_adjacent_cancer_03-115E
Organism Homo sapiens
Characteristics Prostate normal adjacent from cancer patient 03-115E
Extracted molecule total RNA
Extraction protocol Following LCM, captured cells were lysed in RNA extraction buffer (Arcturus). RNA was isolated using the PicoPure RNA Isolation Kit (Arcturus) and the samples were DNAse treated using RNase-Free DNase (Qiagen). Each slide capture session lasted no longer than 20 minutes to minimize RNA degradation. The laser capture settings were: 55 mW beam, 1.5 ms pulse, 15 µm spot size. Subsequently, RNA was amplified through two rounds of linear amplification using the MessageAmp aRNA Kit (Ambion, Inc). Sample quality and quantification was assessed by agarose gel electrophoresis and absorbance at A260.
Label Cy5
Label protocol The protocol used for indirect labeling of cDNAs was a modification of a protocol described elsewhere (http://cmgm.stanford.edu/pbrown/protocols/aadUTPCouplingProcedure.htm). In brief, cDNA probes were made from 2 µg of amplified RNA in a reaction volume of 30 µl containing 5 µg random hexamer primers, 0.2 mM 5-(3-aminoallyl)-2_-deoxyuridine-5_-triphosphate (amino acid-dUTP; Sigma-Aldrich), 0.3 mM dTTP, 0.5 mM each dATP, dCTP, and dGTP, and 380 units of Superscript II reverse transcriptase (Invitrogen) incubated at 42oC for 120 minutes. After RNA hydrolysis, purified cDNA was combined with either Cy3 or Cy5 monoreactive fluors (Amersham) that covalently couple to the cDNA-incorporated aminoallyl linker in the presence of 50 mM NaHCO3 (pH 9.0). Samples were randomly labeled with either Cy3 or Cy5 dye to account for dye-bias. The coupling reaction was quenched with hydroxylamine.
 
Channel 2
Source name Prostate_cancer_03-115E_LCM_GL4
Organism Homo sapiens
Characteristics Prostate Cancer patient 03-115E Gleason_Score:4+3 (tertiary pattern 5)
LCM_Gleason_Pattern:4 Gleason_Pattern:4 Age:70-79 PSA:5.2 Volume:4 Margin_Status:negative Treatment:none
Extracted molecule total RNA
Extraction protocol Following LCM, captured cells were lysed in RNA extraction buffer (Arcturus). RNA was isolated using the PicoPure RNA Isolation Kit (Arcturus) and the samples were DNAse treated using RNase-Free DNase (Qiagen). Each slide capture session lasted no longer than 20 minutes to minimize RNA degradation. The laser capture settings were: 55 mW beam, 1.5 ms pulse, 15 µm spot size. Subsequently, RNA was amplified through two rounds of linear amplification using the MessageAmp aRNA Kit (Ambion, Inc). Sample quality and quantification was assessed by agarose gel electrophoresis and absorbance at A260.
Label Cy3
Label protocol The protocol used for indirect labeling of cDNAs was a modification of a protocol described elsewhere (http://cmgm.stanford.edu/pbrown/protocols/aadUTPCouplingProcedure.htm). In brief, cDNA probes were made from 2 µg of amplified RNA in a reaction volume of 30 µl containing 5 µg random hexamer primers, 0.2 mM 5-(3-aminoallyl)-2_-deoxyuridine-5_-triphosphate (amino acid-dUTP; Sigma-Aldrich), 0.3 mM dTTP, 0.5 mM each dATP, dCTP, and dGTP, and 380 units of Superscript II reverse transcriptase (Invitrogen) incubated at 42oC for 120 minutes. After RNA hydrolysis, purified cDNA was combined with either Cy3 or Cy5 monoreactive fluors (Amersham) that covalently couple to the cDNA-incorporated aminoallyl linker in the presence of 50 mM NaHCO3 (pH 9.0). Samples were randomly labeled with either Cy3 or Cy5 dye to account for dye-bias. The coupling reaction was quenched with hydroxylamine.
 
 
Scan protocol Fluorescent array images were collected for both Cy3 and Cy5 using a GenePix 4000B fluorescent scanner (Axon Instruments, Foster City, CA). The image intensity data were gridded and extracted using GenePix Pro 4.1 software.
Description Human prostate cancer patient 03-115E with Gleason Score 4+3 (tertiary pattern 5) prostate cancer hybridized against normal adjacent prostate tissue from the same patient through laser capture microdissection (LCM).
Data processing Log-Ratios: For each spot and in each channel (Cy3 and Cy5); the median background intensity was subtracted from the median foreground intensity. Log-ratios of cancer expression to benign expression were created by first dividing the background subtracted intensities (CaP/Benign) and then taking the log base 2. If the median background intensity was greater than the median foreground intensity, the spot was considered missing. Removal of Control Genes: The array contained approximately 400 clones used only for quality control purposes (i.e. yeast sequences or blank spots). These clones were removed from the dataset. Lowess Normalization: For each array, the log-ratio data were centered using a print-tip specific Lowess curve (Y. H. Yang, S. Dudoit, P. Luu and T. P. Speed. Normalization for cDNA Microarray Data. SPIE BiOS 2001, San Jose, California, January 2001). This curve was fit to the log intensity versus log-ratio plot using the neighboring 20.0% of the data to calculate the fit at each spot. The Lowess fit at each point was subtracted from the observed log-ratio for that spot, resulting in a normalized log-ratio. Assessing Spot Quality: Spots of poor quality, as determined by both visual inspection and GenePix Pro 4.1 quality flags were considered missing. In addition, spots with background subtracted intensity levels less than 300 were considered missing due to poorly hybridized cDNAs. Clones which were missing on > 20% of arrays were removed from the analysis. Imputation: Missing values were imputed using k-nearest neighbors imputation (k = 10) (2). The dataset was split by Gleason pattern (Grades 3, 4 and 5) and imputation was performed separately for each pattern. Average Replicated Clones: Log-ratios from the replicated cDNA spots on each PEDB chip were averaged after normalization and imputation. These average expression values were used for comparative analysis.
 
Submission date Jun 21, 2006
Last update date Oct 02, 2008
Contact name Denise Mauldin
E-mail(s) dmauldin@fhcrc.org
Phone 2066673480
Fax 2066672917
URL http://www.pedb.org
Organization name Fred Hutchinson Cancer Research Center
Department Human Biology
Lab Peter Nelson
Street address 1100 Fairview Ave N D4-100
City Seattle
State/province WA
ZIP/Postal code 98109
Country USA
 
Platform ID GPL3834
Series (1)
GSE5132 Molecular Correlate to Gleason Grade in Prostate Adenocarcinoma

Data table header descriptions
ID_REF the unique identifier of the feature derived from the Array List
VALUE same as UNF_VALUE but with flagged values removed
X the X-coordinate in µm of the center of the feature-indicator associated with the feature, where (0,0) is the top left of the image.
Y the Y-coordinate in µm of the center of the feature-indicator associated with the feature, where (0,0) is the top left of the image.
Dia. the diameter in µm of the feature-indicator.
F635 Median median feature pixel intensity at wavelength #1 (635 nm).
F635 Mean mean feature pixel intensity at wavelength #1 (635 nm).
F635 SD the standard deviation of the feature pixel intensity at wavelength #1 (635 nm).
B635 Median the median feature background intensity at wavelength #1 (635 nm).
B635 Mean the mean feature background intensity at wavelength #1 (635 nm).
B635 SD the standard deviation of the feature background intensity at wavelength #1 (635 nm).
% > B635+1SD the percentage of feature pixels with intensities more than one standard deviation above the background pixel intensity, at wavelength #1 (635 nm).
% > B635+2SD the percentage of feature pixels with intensities more than two standard deviations above the background pixel intensity, at wavelength #1 (635 nm).
F635 % Sat. the percentage of feature pixels at wavelength 635 that are saturated.
F532 Median median feature pixel intensity at wavelength #2 (532 nm).
F532 Mean mean feature pixel intensity at wavelength #2 (532 nm).
F532 SD the standard deviation of the feature intensity at wavelength #2 (532 nm).
B532 Median the median feature background intensity at wavelength #2 (532 nm).
B532 Mean the mean feature background intensity at wavelength #2 (532 nm).
B532 SD the standard deviation of the feature background intensity at wavelength #2 (532 nm).
% > B532+1SD the percentage of feature pixels with intensities more than one standard deviation above the background pixel intensity, at wavelength #2 (532 nm).
% > B532+2SD the percentage of feature pixels with intensities more than two standard deviations above the background pixel intensity, at wavelength #2 (532 nm).
F532 % Sat. the percentage of feature pixels at wavelength 532 that are saturated.
Ratio of Medians (635/532) the ratio of the median intensities of each feature for each wavelength, with the median background subtracted.
Ratio of Means (635/532) the ratio of the arithmetic mean intensities of each feature for each wavelength, with the median background subtracted.
Median of Ratios (635/532) the median of pixel-by-pixel ratios of pixel intensities, with the median background subtracted.
Mean of Ratios (635/532) the geometric mean of the pixel-by-pixel ratios of pixel intensities, with the median background subtracted.
Ratios SD (635/532) the geometric standard deviation of the pixel intensity ratios.
Rgn Ratio (635/532) the regression ratio of every pixel in a 2-feature-diameter circle around the center of the feature.
Rgn R² (635/532) the coefficient of determination for the current regression value.
F Pixels the total number of feature pixels.
B Pixels the total number of background pixels.
Sum of Medians the sum of the median intensities for each wavelength, with the median background subtracted.
Sum of Means the sum of the arithmetic mean intensities for each wavelength, with the median background subtracted.
Log Ratio (635/532) log (base 2) transform of the ratio of the medians.
F635 Median - B635 the median feature pixel intensity at 635 with the median background subtracted.
F532 Median - B532 the median feature pixel intensity at 532 with the median background subtracted.
F635 Mean - B635 the mean feature pixel intensity at 635 with the median background subtracted.
F532 Mean - B532 the mean feature pixel intensity at 532 with the median background subtracted.
F635 Total Intensity the sum of all pixel intensities in the feature at wavelenth 635.
F532 Total Intensity the sum of all pixel intensities in the feature at wavelenth 532.
SNR 635 the signal-to-noise ratio of the feature, calculated as (F635 Mean - B635 Mean)/B635 SD.
SNR 532 the signal-to-noise ratio of the feature, calculated as (F532 Mean - B532 Mean)/B532 SD.
Flags the type of flag associated with a feature; GenePix Pro 4.1 quality flags 0, -50, -75
Normalize the normalization status of the feature (included/not included).
INV_VALUE
UNF_VALUE -[INV_VALUE]; Lowess-normalized, log2(CaP/Benign) ratios

Data table
ID_REF VALUE X Y Dia. F635 Median F635 Mean F635 SD B635 Median B635 Mean B635 SD % > B635+1SD % > B635+2SD F635 % Sat. F532 Median F532 Mean F532 SD B532 Median B532 Mean B532 SD % > B532+1SD % > B532+2SD F532 % Sat. Ratio of Medians (635/532) Ratio of Means (635/532) Median of Ratios (635/532) Mean of Ratios (635/532) Ratios SD (635/532) Rgn Ratio (635/532) Rgn R² (635/532) F Pixels B Pixels Sum of Medians Sum of Means Log Ratio (635/532) F635 Median - B635 F532 Median - B532 F635 Mean - B635 F532 Mean - B532 F635 Total Intensity F532 Total Intensity SNR 635 SNR 532 Flags Normalize INV_VALUE UNF_VALUE
89663 -1.83695 2290 14030 130 4338 5017 2294 127 142 85 100 100 0 1345 1555 733 196 204 61 100 99 0 3.665 3.598 3.5 3.76 1.428 3.824 0.783 120 709 5360 6249 1.874 4211 1149 4890 1359 602035 186571 57.353 22.148 0 0 1.836949878 -1.83695
89664 1.01581 2480 14030 130 13118 13008 3206 130 223 1013 100 100 0 28727 28299 6829 207 384 1874 100 100 0 0.455 0.458 0.455 0.459 1.207 0.434 0.924 120 638 41508 40970 -1.135 12988 28520 12878 28092 1561002 3395898 12.621 14.896 0 0 -1.015807226 1.01581
89665 1.79125 2670 14030 130 2842 3151 1614 125 133 60 100 100 0 8827 9791 4305 196 209 93 100 100 0 0.315 0.315 0.294 0.306 1.356 0.296 0.823 120 627 11348 12621 -1.668 2717 8631 3026 9595 378163 1174911 50.3 103.032 0 0 -1.791251145 1.79125
89666 -0.0674656 2870 14020 130 3079 3302 1172 121 127 48 100 100 0 2878 3212 1224 191 196 44 100 100 0 1.101 1.053 1.05 1.068 1.281 1.014 0.844 120 633 5645 6202 0.139 2958 2687 3181 3021 396291 385477 66.146 68.545 0 0 0.067465571 -0.0674656
89667 -0.0934342 3060 14030 130 3356 3401 1029 124 153 371 100 99 0 3062 2988 1020 194 199 45 100 100 0 1.127 1.173 1.152 1.208 1.391 1.15 0.864 120 609 6100 6071 0.172 3232 2868 3277 2794 408176 358571 8.755 61.978 0 0 0.093434192 -0.0934342
89668 -3.78333 3260 14040 130 21140 20746 4350 126 156 368 100 100 0 1596 1635 421 193 198 43 100 100 0 14.978 14.3 14.751 14.643 1.277 14.558 0.89 120 620 22417 22062 3.905 21014 1403 20620 1442 2489505 196218 55.951 33.419 0 0 3.783334629 -3.78333
89669 -0.148207 3450 14020 130 22351 22159 5744 125 139 86 100 100 0 22357 21947 6674 196 215 285 100 100 0 1.003 1.013 0.98 1.053 1.515 0.961 0.87 120 696 44387 43785 0.004 22226 22161 22034 21751 2659087 2633583 256.047 76.253 0 0 0.148207303 -0.148207
89670 -3.61435 3660 14020 130 6712 6727 1575 123 135 76 100 100 0 728 803 309 196 207 80 99 99 0 12.385 10.88 12.374 11.947 1.471 10.354 0.725 120 616 7121 7211 3.631 6589 532 6604 607 807213 96397 86.737 7.45 0 0 3.614352345 -3.61435
89671 -0.0365511 3850 14020 140 20967 19866 5479 122 127 39 100 100 0 22573 21347 6328 192 196 49 100 100 0 0.931 0.933 0.921 0.951 1.417 0.917 0.925 156 640 43226 40899 -0.103 20845 22381 19744 21155 3099164 3330186 506.128 431.653 0 0 0.036551087 -0.0365511
89672 0.803676 4030 14030 130 11755 11813 2262 123 147 178 100 100 0 21753 21383 5107 193 273 902 100 100 0 0.54 0.552 0.528 0.563 1.314 0.518 0.918 120 582 33192 32880 -0.89 11632 21560 11690 21190 1417607 2565913 65.539 23.404 0 0 -0.803675952 0.803676
89673 -0.445447 4250 14020 130 4117 4130 1175 121 157 316 100 100 0 2956 2909 746 192 347 1763 90 15 0 1.446 1.476 1.492 1.462 1.31 1.566 0.893 120 652 6760 6726 0.532 3996 2764 4009 2717 495551 349029 12.573 1.453 0 0 0.445446918 -0.445447
89674 -0.522587 4430 14020 140 1194 1246 367 118 135 265 98 98 0 965 1073 449 187 272 1519 7 0 0 1.383 1.273 1.355 1.322 1.457 1.168 0.771 156 677 1854 2014 0.468 1076 778 1128 886 194410 167461 4.192 0.527 0 0 0.522586968 -0.522587
89675 0.470546 4630 14020 140 6365 6169 2000 117 132 75 99 99 0 8243 8229 2331 188 198 64 100 100 0 0.776 0.753 0.751 0.708 1.553 0.752 0.904 156 762 14303 14093 -0.366 6248 8055 6052 8041 962303 1283778 80.493 125.484 0 0 -0.470545938 0.470546
89676 -0.320582 4850 14030 90 2680 2996 1872 124 163 118 100 98 0 2186 2616 1662 195 215 91 100 100 0 1.284 1.186 1.333 1.253 1.847 1.168 0.837 52 352 4547 5293 0.36 2556 1991 2872 2421 155779 136049 24.008 26.385 0 0 0.320582178 -0.320582
89677 -2.68283 5010 14010 130 13033 12675 2570 122 139 148 100 100 0 2036 1937 406 190 201 90 100 100 0 6.994 7.185 6.904 7.306 1.327 7.419 0.909 120 647 14757 14300 2.806 12911 1846 12553 1747 1520953 232456 84.703 19.289 0 0 2.682834031 -2.68283
89678 -0.158923 5220 14010 140 4716 4795 1669 117 130 135 100 100 0 3990 4162 1292 190 200 86 100 100 0 1.21 1.178 1.152 1.144 1.423 1.201 0.891 156 705 8399 8650 0.275 4599 3800 4678 3972 748082 649220 34.556 46.07 0 0 0.158923024 -0.158923
89679 -0.330137 5410 14020 130 1877 2297 1542 117 123 42 100 100 0 1556 1994 1350 192 198 50 100 100 0 1.29 1.21 1.189 1.215 1.462 1.207 0.843 120 555 3124 3982 0.368 1760 1364 2180 1802 275657 239309 51.762 35.92 0 0 0.330136795 -0.330137
89680 0.536682 5600 14010 140 9921 10522 3353 115 120 37 100 100 0 14762 14653 5532 189 194 46 99 99 0 0.673 0.72 0.709 0.823 2.079 0.678 0.895 156 669 24379 24871 -0.572 9806 14573 10407 14464 1641390 2285846 281.135 314.326 0 0 -0.53668155 0.536682
89681 0.634624 5800 14010 140 2409 2465 851 115 121 44 100 100 0 3580 3746 1224 187 202 133 100 100 0 0.676 0.66 0.654 0.654 1.394 0.639 0.859 156 667 5687 5909 -0.565 2294 3393 2350 3559 384597 584452 53.273 26.647 0 0 -0.634623661 0.634624
89682 0.539499 5990 14020 140 1972 1907 596 113 119 47 100 100 0 2819 2730 863 186 196 97 100 100 0 0.706 0.705 0.681 0.711 1.427 0.684 0.869 156 637 4492 4338 -0.502 1859 2633 1794 2544 297558 425831 38.043 26.124 0 0 -0.539498613 0.539499

Total number of rows: 15488

Table truncated, full table size 3562 Kbytes.




Supplementary file Size Download File type/resource
GSM115773.gpr.gz 1.3 Mb (ftp)(http) GPR

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