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Sample GSM115767 Query DataSets for GSM115767
Status Public on Jul 14, 2006
Title Prostate_03-159_GL4
Sample type RNA
 
Channel 1
Source name Prostate_cancer_03-159_LCM_GL4
Organism Homo sapiens
Characteristics Prostate Cancer patient 03-159 Gleason_Score:3+4 LCM_Gleason_Pattern:4 Gleason_Pattern:4 Age:60-69 PSA:4.7 Volume:3 Margin_Status:positive 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 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_normal_adjacent_cancer_03-159
Organism Homo sapiens
Characteristics Prostate normal adjacent from cancer patient 03-159
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 03-159 with Gleason Score 3+4 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).
UNF_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 UNF_VALUE
89663 -0.933351167 2430 14610 110 478 553 265 96 101 44 100 100 0 1157 1410 620 100 104 51 100 100 0 0.361 0.349 0.325 0.328 1.479 0.326 0.77 80 571 1439 1767 -1.468 382 1057 457 1310 44218 112820 10.273 25.608 0 0 -0.933351167
89664 1.753634084 2620 14620 120 6856 6669 1394 97 104 47 100 100 0 2597 2516 586 103 108 54 98 98 0 2.71 2.724 2.621 2.851 1.392 2.697 0.939 120 635 9253 8985 1.438 6759 2494 6572 2413 800253 301932 139.681 44.593 0 0 1.753634084
89665 1.246018403 2820 14610 110 1270 1497 678 97 106 61 100 100 0 807 946 403 101 106 48 100 100 0 1.661 1.657 1.625 1.654 1.241 1.777 0.891 80 560 1879 2245 0.732 1173 706 1400 845 119730 75700 22.803 17.5 0 0 1.246018403
89666 0.771461369 3030 14620 120 1460 1479 424 99 106 59 100 100 0 1213 1196 348 100 105 45 100 100 0 1.223 1.259 1.307 1.261 1.335 1.306 0.872 120 632 2474 2476 0.29 1361 1113 1380 1096 177499 143545 23.271 24.244 0 0 0.771461369
89667 0.768433702 3210 14620 110 1372 1405 355 102 112 78 100 100 0 1149 1249 364 104 116 120 100 100 0 1.215 1.138 1.158 1.149 1.292 1.084 0.875 80 506 2315 2448 0.281 1270 1045 1303 1145 112389 99932 16.577 9.442 0 0 0.768433702
89668 -0.609016026 3420 14610 120 1306 1301 347 103 113 72 100 99 0 2593 2582 443 105 115 110 100 100 0 0.484 0.484 0.492 0.461 1.372 0.487 0.916 120 614 3691 3675 -1.048 1203 2488 1198 2477 156173 309808 16.5 22.427 0 0 -0.609016026
89669 0.709505059 3600 14610 120 8582 8485 1787 103 110 46 100 100 0 5770 5652 1177 105 107 26 100 100 0 1.497 1.511 1.519 1.521 1.194 1.519 0.946 120 569 14144 13929 0.582 8479 5665 8382 5547 1018160 678263 182.065 213.269 0 0 0.709505059
89670 -0.558223345 3800 14610 120 351 364 118 107 112 46 100 97 0 596 596 109 104 107 23 100 100 0 0.496 0.522 0.496 0.482 1.536 0.519 0.686 120 599 736 749 -1.012 244 492 257 492 43659 71495 5.478 21.261 0 0 -0.558223345
89671 1.221382434 4000 14620 120 11069 10999 1913 105 108 45 100 100 0 5139 5197 1012 101 103 20 100 100 0 2.176 2.138 2.113 2.168 1.188 2.127 0.96 120 579 16002 15990 1.122 10964 5038 10894 5096 1319821 623604 242.022 254.7 0 0 1.221382434
89672 1.081351803 4190 14610 120 6514 6437 923 103 110 48 100 100 0 3746 3664 549 102 104 19 100 100 0 1.759 1.778 1.769 1.801 1.206 1.773 0.956 120 585 10055 9896 0.815 6411 3644 6334 3562 772394 439723 131.813 187.368 0 0 1.081351803
89673 0.303138167 4380 14610 120 959 970 174 109 115 52 100 100 0 1089 1099 171 106 109 33 100 100 0 0.865 0.867 0.86 0.865 1.268 0.853 0.891 120 584 1833 1854 -0.21 850 983 861 993 116392 131886 16.442 30 0 0 0.303138167
89674 -0.481648668 4580 14610 120 321 335 101 109 113 47 97 92 0 513 520 74 105 107 23 100 100 0 0.52 0.545 0.54 0.487 1.839 0.567 0.627 120 599 620 641 -0.945 212 408 226 415 40192 62346 4.723 17.957 0 0 -0.481648668
89675 -0.576106205 4780 14600 120 1883 1878 356 97 105 49 100 100 0 3719 3755 686 98 104 30 100 100 0 0.493 0.487 0.49 0.485 1.19 0.485 0.931 120 693 5407 5438 -1.02 1786 3621 1781 3657 225325 450617 36.184 121.7 0 0 -0.576106205
89676 -0.051399557 5000 14620 70 589 668 316 108 122 67 96 96 0 832 789 346 108 123 58 100 100 0 0.664 0.822 0.854 0.797 1.7 0.863 0.806 32 231 1205 1241 -0.59 481 724 560 681 21391 25238 8.149 11.483 0 0 -0.051399557
89677 0.070158778 5170 14610 120 1558 1620 339 98 104 44 100 100 0 1988 2019 313 99 102 26 100 100 0 0.773 0.793 0.783 0.784 1.172 0.793 0.921 120 630 3349 3442 -0.372 1460 1889 1522 1920 194356 242272 34.455 73.731 0 0 0.070158778
89678 0.490588936 5370 14610 130 1347 1394 354 99 102 41 100 100 0 1334 1390 284 95 96 17 100 100 0 1.007 1 1.01 0.983 1.237 1.069 0.9 120 662 2487 2590 0.01 1248 1239 1295 1295 167310 166850 31.512 76.118 0 0 0.490588936
89679 0.96143126 5560 14610 120 1367 1614 776 103 106 45 100 100 0 1013 1182 576 99 102 21 100 99 0 1.383 1.395 1.393 1.436 1.371 1.403 0.923 120 584 2178 2594 0.468 1264 914 1511 1083 193659 141783 33.511 51.429 0 0 0.96143126
89680 1.480035368 5760 14610 120 8142 8022 1584 101 107 46 100 100 0 3500 3438 595 97 100 20 100 100 0 2.363 2.371 2.361 2.365 1.176 2.439 0.948 120 598 11444 11262 1.241 8041 3403 7921 3341 962638 412573 172.065 166.9 0 0 1.480035368
89681 0.494324549 5950 14600 130 1895 1917 316 90 97 43 100 100 0 1826 1865 258 89 90 16 100 100 0 1.039 1.029 1.017 1.026 1.181 1.037 0.923 120 669 3542 3603 0.055 1805 1737 1827 1776 230074 223845 42.326 110.938 0 0 0.494324549
89682 0.510001379 6140 14600 120 1170 1213 222 94 98 43 100 100 0 1156 1157 158 92 93 16 100 100 0 1.011 1.051 1.051 1.043 1.188 1.057 0.917 120 570 2140 2184 0.016 1076 1064 1119 1065 145564 138810 25.93 66.5 0 0 0.510001379

Total number of rows: 15488

Table truncated, full table size 3335 Kbytes.




Supplementary file Size Download File type/resource
GSM115767.gpr.gz 1.2 Mb (ftp)(http) GPR

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