title: Pregnant females fed with maltose dextrin, PF07 (Pair Fed) corresponds to replicate 7

status: Public on Oct 31 2006

description: Timed pregnant C57Bl6 mice were exposed to ethanol during gestational day 15 to day 18 and ethanol exposed fetuses were delivered by cessarian section on gestational day 19. Alcohol exposed fetuses were compared to control timed preg

hyb_protocol: Hybridization was performed according to Genisphere's 3DNA Array 900 kit

scan_protocol: Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000B fluorescent scanner and image intensity data were extracted and analyzed with GenePix Pro 6.0.1.26 analysis software

data_processing: Background correction and removal of flagged values were conducted prior to any further analysis

source_name: Pair Fed 07

label_protocol: 1ug of total RNA reverse-transcribed and labeled according Genisphere's 3DNA Array 900 Cy3/Cy5 kit's protocol

growth_protocol: Time pregnant C57Bl6 mice were fed adlibitum with food and water prior to feeding with maltose dextrin (corresponding to gestational day 14)

extract_protocol: Total RNA extracted with Ambion's miRNA isolation kit

treatment_protocol: Timed pregnant C57Bl6 mice fed with maltose dextrin from gestational day 15 to 18 (pair fed group)

tag: Total RNA extracted from the cerebral cortex of timed pregnant C57B16 mice (pair fed group)

source_name: Weight matched control

label_protocol: A total of 1ug of total RNA was reverse-transcribed with Superscript II (Invitrogen) using primers for Cy3 and Cy5 according to the 3DNA Array 900 Cy3/Cy5 protocol (Genisphere).

growth_protocol: Time pregnant C57Bl6 mice were fed adlibitum with food and water prior to the experiment and during the experiment

treatment_protocol: Pregnant animals were fed adlibitum with food and water (control)

tag: Total RNA extracted from the cerebral cortex of 7 animals were combined and to be used as a normalization control


title: Pregnant females fed with maltose dextrin, PF06 (Pair Fed) corresponds to replicate 6

type: RNA

city: Bryan

first: Rajesh

last: Miranda

address: 8447 Riverside Parkway

country: USA

institute: Texas A&M University

molecule: total RNA

organism: Mus musculus

source_name: Pair Fed 06


title: Pregnant females fed with maltose dextrin, PF05 (Pair Fed) corresponds to replicate 5

source_name: Pair Fed 05


title: Mouse cDNA microarray analysis of CHB50 cells, 0226BESS corresponds to replicate 2

status: Public on Dec 01 2006

description: To identify neuronal differentiation-associated gene candidates using CHB50 neuroblasts by comparing CHB50 cells from the permissive condition (proliferation) to the non-permissive condition (differentiation)

data_processing: After background correction and removal of flagged values (including controls and housekeeping genes) the log ratio (VALUE) was calculated by Ch1 (P1Signal) divided by Ch2 (P2BalancedSignal)

molecule: polyA RNA

organism: Rattus norvegicus

source_name: CHB50 at 33C (permissive temperature/+tsTA/-p53))

growth_protocol: CHB50 cultures are maintained in DMEM, 10%Fetal Calf Serum, 100U/ml penicillin-streptomycin and 50ug/ml of G418

extract_protocol: Trizol method (Invitrogen) followed by poly-A-mRNA isolation with Qiagen's Oligotex mRNA kit

treatment_protocol: CHB50 cells cultured at the permissive temperature (33C)

tag: Conditionally immortalized CHB50 cells are cultured at 33C for expression of the large T antigen and attenuation of p53 (the proliferation condition)

source_name: CHB50 at 39C (non-permissive temperature/-tsTA/+p53)

growth_protocol: CHB50 cultures are maintained in serum-free medium in DMEM, 49%F-12, 1%N2 supplement, and 100U/ml penicillin-streptomycin

treatment_protocol: CHB50 cells cultured at the non-permissive temperature (39C) for 72 hours

tag: Conditionally immortalized CHB50 cells are cultured at 39C for suppression of the large T antigen and induction of p53 (the differentiation condition)


title: B129.2 RA TNF t=0 PAXgene vs CRS red 80 PMT

status: Public on Jan 28 2010

city: Amsterdam

first: Lisa

last: van Baarsen

address: Meibergdreef 9

department: Div. of Clinical Immunology and Rheumatology

country: Netherlands

institute: EULAR & FOCIS Center of Excellence

description:
Simple annotation: Normal tissue, Whole blood
Image: http://smd.stanford.edu/MicroArray/gifs/2004-07/53847.gif

hyb_protocol: not provided

scan_protocol: GenePix

data_processing: VALUE is Log (base 2) of the ratio of the median of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm)

organism: Homo sapiens

source_name: B129.2 RA TNF t=0 PAX

label_protocol: Indirect labelling of amplified RNA via aminoallyl; As described in: Gene expression profiling in rheumatology. Methods Mol Med. 2007;136:305-27  The following is a slight modification by Mitch Garber, Anatoly Urisman and subsequently by Lisa van Baarsen of a protocol developed by Joe DeRisi (UCSF) and Rosetta Inpharmatics (Kirkland, WA).  Original document can be obtained at www.microarrays.org.   cDNA synthesis:  Two separate reaction vials for labeling: one for the experimental sample to be labeled with Cy3 (green) and one for the reference sample to be labeled with Cy5 (red).    1. For total RNA, mix 25 ug of the RNA with 5 ug of anchored oligo-dT [(dT)20-     VN] (Operon, HPLC purified) in a  total volume of 18 ul in 1,5 ml tubes. For   cRNA, use 5 ug of the amplified cRNA with 10 ug of Poly D(N)6 (Operon) primer   in a total volume of 18 ul in 1.5 ml tubes.  2. Heat to 70C for 10 minutes at 300 rpm  3. Cool on ice water for 5 minutes. 4. Make a nucleotide master mix  Nucleotide Mix per reaction: 6 ul 5X RT buffer 0.6 ul 50X dNTP stock solution* 3 ul 0.1 M DTT 1.5 ul 200 U/ul Superscript II RT (Gibco) 0.5 ul RNasin (Gibco)    *50X dNTP stock solution using a 4:1 ratio aminoallyl-dUTP to dTTP  10 ul each 100 mM dATP, dGTP, dCTP (Pharmacia)  8 ul 100 mM aminoallyl-dUTP** (Sigma, #A0410)  2 ul 100 mM dTTP **Dissolve 10 mg aminoallyl-dUTP in 170 uL water.  Add approx. 6.8 uL 1N NaOH.     Final pH is roughly 7.0 using pH paper.  5. Pipet 11.6 ul nucleotide mix in a 0.5 ml thin wall tube for both the Cy3 and Cy5 samples 4. Spin down and add the 18 ul of sample or reference  5. Incubate reaction for 1h at 42C 6. Add 1 ul reverse transcriptase and continue incubation at 42C for an additional 1h   Hydrolysis of RNA: 7. Add 15 ul of 0.1 N NaOH.  Incubate at 70C for 10 minutes if cRNA is used or 30 minutes if total RNA is used as template  8. Add 15 ul 0.1 N HCl.    9. Carefully add 450 ul RNAse-free water to each reaction.   Cleanup: 10. Apply the 500 ul neutralized, diluted reaction mix on a labeled Microcon-30 filter (Amicon), spin at 10,000g for 7 minutes and discard flow through 11. Pipet 450 ul water on the filter, spin at 10,000g for 7 minutes and discard flow through 12. Repeat washing step 11 13. Concentrate the probe to 10 ul by centrifugation in steps of 30 seconds. Check after each step the fluid level in the filter. Never let the membrane dry completely!! 14. If there is less then half a circle of fluid on top of the filter, invert the microcon in a clean labeled tube and spin at 10,000g for 1 minute to recover the aminoallyl labeled cDNA probe; adjust to 10 ul with water 15. Samples can now be stored at -20C or -80C   Coupling of aminoallyl-labeled cDNA to Cy3 or Cy5 16. Add 0.5 ul 1M sodium bicarbonate, pH 9.0 (50 mM final) to the 10 ul aminoallyl-labeled cDNA and transfer the sample to the right aliquot of dye () 17. Incubate 1h at RT in the dark.  Mix every 15 minutes.   Quenching and Cleanup 18. Add 4.5 ul 4M hydroxylamine  19. Incubate 15 minutes in the dark    To remove unincorporated/quenched Cy dyes, proceed with Qia-Quick PCR purification kit (QIAGEN). Use method as described by the manufacturer.   Use Cy3/5 labeled samples immediately for hybridisation; Protocol Type = Labeling; Performer: Cornelis,,Verweij

extract_protocol: Linear amplification of total RNA using Ambion kit; As described in: Gene expression profiling in rheumatology. van der Pouw Kraan TC, van Baarsen LG, Rustenburg F, Baltus B, Fero M, Verweij CL. Methods Mol Med. 2007;136:305-27  Protocol MessageAmp aRNA kit (Ambion #1750; version 0309A; updated 27-04-2004)  All incubation steps are performed in a thermal cycler block using thin wall tubes. When the incubation temperature is ??37??C a heated lid is used. Do not use a heated lid when incubation temperature is <37??C. Use as input 1 microgram (ug) total RNA in a maximal volume of 11 ul.  First strand synthesis: 1. Thaw samples and reagents 2. Set the temperature of the thermal cycler on 70 degrees Celcius (C)  3. Prepare the following mix:     - RNA sample (max 11 ul)        - 1 ul oligo T7 primer         - add RNAse free water to a final volume of 12 ul  4. Incubate samples 10 minutes at 70C in thermal cycler 5. Put on ice-water for 5 minutes 6. Prepare Reverse transcriptase mix at RT, use per reaction:     2 ul 10x First strand buffer          1 ul Ribonuclease inhibitor          4 ul dNTP mix    1 ul Reverse Transcriptase    7. Add 8 ul mastermix to each sample 8. Incubate for 2h at 42??C   Second strand synthesis: 9. Prepare second strand (SS) mix, use per reaction:        63 ul RNAse free water          10 ul 10x second strand buffer          4 ul dNTP mix            2 ul DNA polymerase                1 ul RNAse H                         10. Add 80 ul SS mix to each sample  11. Incubate for 2h at 16??C 12. (If desired; samples can be immediately stored at ?C20C until the next day)  cDNA Purification (fume hood): 13. Pre-heat RNAse free water at 50??C 14. Take 1 tube en 1 cDNA filter per cDNA sample 15. Add 50 ul cDNA binding buffer to the filter and incubate for 5 minutes at RT 16. Pipet 250 ul DNA binding buffer to the cDNA samples 17. Pipet cDNA sample mix on the pre-wetted filter, centrifuge 1 min at 10.000g and discard flowthrough  18. Add 500 ul cDNA wash buffer to the filter and centrifuge 1 min at 10.000g, discard flow through 19. Centrifuge another 1 min at 10.000g  20. Put filter in a clean 2 ml elution tube 21. Pipet 10 ul pre-heated (50C) RNAse free water on the filter, incubate 2 min at RT and centrifuge 2 min at 10.000 rpm 22. Repeat step 21 23. Discard filter 24. Measure in each sample the volume; should be 16-18 ul adjust if necessary 25. Store cDNA at ?C20C until the next day or at ?C80C for a longer time period)  In vitro transcription to synthesize aRNA 1. Set the temperature of the thermal cycler on 37C  2. Thaw the reagents needed for the in vitro transcription on ice and put them at RT once they are thawed 3. Prepare at RT the mix containing per sample:     4 ul of each ATP, CTP, GTP nucleotide vial (75 mM)     4 ul UTP Solution (75 mM)    4 ul T7 10x reaction buffer                 4 ul T7 Enzyme mix               4. Pipet 24 ul mix to the samples 5. Incubate for 5h at 37??C 6. Put the samples on ice  7. Add 2 ul DNAse I incubate for 30 min at 37C  aRNA Purification (fume hood RT) 8. Pre-heat 100 ul RNAse free water per sample at 50??C  9. Take 1 clean tube and 1 filter per cRNA sample  10. Pipet 100 ul RNA binding buffer on the filter and incubate for 5 min  11. Pipet 58 ul RNA elution solution to each cRNA sample, vortex and add 350 ul cRNA binding buffer to each sample and vortex 12. Transfer samples to a new 1.5 ml tube and add 250 ul 100% ethanol  13. Apply the 650 ul mix on the pre-wetted filter, spin 1 min at 10.000g and discard flow through 14. Add 650 ul aRNA wash buffer to the filter, spin 1 min at 10.000g  15. Spin 1 min at 10.000g and move the dry filter to a fresh 2 ml tube  16. Add 50 ul pre-heated RNAse free water, incubate 2 min  17. Spin 2 min at 10.000g and repeat step 16 18. Spin 2 min at 10.000 rpm and discard filter  19. Measure the quantity and quality of cRNA on Agilent BioAnalyzer or NanoDrop ND-1000 spectrophotometer and store at ?C80??C; Protocol Type = Amplification; Performer: Cornelis,,Verweij

tag: patient

tag: time(absolute)

value: 0 months

tag: clinical (delta das28)

tag: clinical (eular)

tag: clinical (ddasv2-v10)

label_protocol: not provided

extract_protocol: not provided

tag: reference


title: Pregnant females fed with maltose dextrin, PF01 (Pair Fed) corresponds to replicate 1

source_name: Pair Fed 01


title: Pregnant females fed with maltose dextrin, PF04 (Pair Fed) corresponds to replicate 4

source_name: Pair Fed 04


title: B142 RA TNF t=0 PAXgene vs CRS red 80 PMT

description:
Simple annotation: Normal tissue, Whole blood
Image: http://smd.stanford.edu/MicroArray/gifs/2004-06/53087.gif

source_name: B142 RA TNF t=0 PAX


title: Pregnant females exposed to ethanol for 5 days, AF06 (Alcohol Fed) corresponds to replicate 6

source_name: Alcohol Fed 06

growth_protocol: Time pregnant C57Bl6 mice were fed adlibitum with food and water prior to ethanol exposure (corresponding to gestational day 14)

treatment_protocol: Timed pregnant C57Bl6 mice exposed to about 180-200mg/dl ethanol for 4 days corresponding to gestation day 15 to 18

tag: Total RNA extracted from the cerebral cortex of timed pregnant C57B16 mice exposed to alcohol


title: Mouse neurospheres cultured for 108 hours, Prominin_03 corresponds to replicate 3

status: Public on Oct 01 2006

hyb_protocol: Hybridization was performed as stated in Genisphere's 3DNA Array 900 manual

scan_protocol: Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000B fluorescent scanner and image intensity data were extracted and analyzed with GenePix Pro 4.1.1.21 analysis software

data_processing: After background correction and removal of flagged values

label_protocol: cDNA was prepared from 1 ug total RNA using Superscript II RNase  H Reverse Transcriptase and a 3DNA dendrimer specific primer was used to capture the sequence for the dendrimer conjugated to Cy5

growth_protocol: Neurospheres were cultured in serum-free mitogenic medium DMEM/F-12, 20ng/ml basic Fibroblast Growth Factor, 20ng/ml human recombinant Epidermal Growth Factor, 0.15ng/ml human recombinant Leukemia Inhibitory Factor, 1X Insuli

extract_protocol: Invitrogen's Trizol method followed by purification with Qiagen's RNeasy kit

treatment_protocol: Neurospheres cultured in growth media and an adenovirus expressing CD133/R-prominin1 for 108hrs (4.5days)

tag: Gestational Day 12.5 Mouse cortical-derived neurospheres that overexpress CD133/R-prominin1

source_name: Universal control embryonic RNA

label_protocol: 1ug of universal control whole embryo RNA was reverse transcribed with a primer containing a sequence complementary to the capture sequence for the dendrimer conjugated to Cy3

growth_protocol: Whole embryos were obtained from timed-pregnant C57/Bl6 mice at gestational day 12

treatment_protocol: Total RNA was extracted from ED12 mouse whole embryos

tag: Universal control embryonic RNA pooled from ED12 mouse whole embryos, used as a normalization control


title: Pregnant females fed with maltose dextrin, PF03 (Pair Fed) corresponds to replicate 3

source_name: Pair Fed 03


title: Pregnant females exposed to ethanol for 5 days, AF03 (Alcohol Fed) corresponds to replicate 3

description: Timed pregnant C57Bl6 mice were exposed to ethanol during gestational day 15 to day 18 and ethanol exposed fetuses were delivered by cessarian section on gestational day 19. Alcohol exposed fetuses were compared to control timed pregnant mice and to pair fed controls (animals fed with maltose dextrose)

source_name: Alcohol Fed 03


title: Pregnant females fed with maltose dextrin, PF02 (Pair Fed) corresponds to replicate 2

source_name: Pair Fed 02


title: Pregnant females exposed to ethanol for 5 days, AF02 (Alcohol Fed) corresponds to replicate 2

source_name: Alcohol Fed 02


title: Mouse neurospheres cultured for 24 hours, Prominin_01 corresponds to replicate 1

description: Gestational Day 12.5 Mouse cortical-derived neurospheres


title: Mouse neurospheres cultured for 24 hours, Prominin_02 corresponds to replicate 2


title: Mouse neurospheres cultured for 24 hours, Prominin_03 corresponds to replicate 3


title: Pregnant females exposed to ethanol for 5 days, AF07 (Alcohol Fed) corresponds to replicate 7

source_name: Alcohol Fed 07


title: Mouse neurospheres cultured for 108 hours, Prominin_01 corresponds to replicate 1


title: Pregnant females exposed to ethanol for 5 days, AF04 (Alcohol Fed) corresponds to replicate 4

source_name: Alcohol Fed 04


title: Mouse neurospheres cultured for 108 hours, Control_01 corresponds to replicate 1

treatment_protocol: Neurospheres cultured in growth media only

tag: Differentiated gestational Day 12.5 Mouse cortical-derived neurospheres (control)


title: Mouse neurospheres cultured for 108 hours, Prominin_02 corresponds to replicate 2


title: Mouse neurospheres cultured for 24 hours, Control_02 corresponds to replicate 2

tag: Gestational Day 12.5 Mouse cortical-derived neurospheres (control)


title: Mouse neurospheres cultured for 24 hours, Control_01 corresponds to replicate 1


title: Mouse neurospheres cultured for 24 hours, Control_03 corresponds to replicate 3


title: Pregnant females exposed to ethanol for 5 days, AF05 (Alcohol Fed) corresponds to replicate 5

source_name: Alcohol Fed 05

source_name: Weight matched control


title: Mouse neurospheres cultured for 108 hours, Control_03 corresponds to replicate 3


title: Mouse neurospheres cultured for 108 hours, Control_02 corresponds to replicate 2


title: Pregnant females exposed to ethanol for 5 days, AF01 (Alcohol Fed) corresponds to replicate 1

source_name: Alcohol Fed 01


title: Mouse neurospheres differentiated in the presence of FGF, b4 corresponds to replicate 6

status: Public on Sep 30 2006

scan_protocol: Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000B fluorescent scanner and image intensity data were extracted and analyzed with GenePix Pro 6.0.1.25 analysis software

data_processing: After background correction and removal of flagged values, background corrected intensities were normalized to the mean-of-ratios

label_protocol: labeled with Genisphere's 3DNA Array 900 Cy3/Cy5 kit

growth_protocol: Neurospheres were cultured in serum-free mitogenic medium DMEM/F-12, 20ng/ml basic Fibroblast Growth Factor, 20ng/ml human recombinant Epidermal Growth Factor, 0.15ng/ml human recombinant Leukemia Inhibitory Factor, 1X Insulin-Transferrin-Selenium-X Supplement, 5ug/ml heparin, and 20nM progesterone and cultured at 37?C (5% CO2) in a humidified chamber to allow formation of neurospheres

treatment_protocol: Neurospheres differentiated by the removal of EGF and LIF from the mitogenic media (control)

label_protocol: A total of 1ug of total RNA was reverse-transcribed with Superscript II (Invitrogen) using primers for Cy3 and Cy5 according to the 3DNA Array 900 Cy3/Cy5 protocol (Genisphere)

growth_protocol: Neurospheres were grown in mitogenic media and supplemented with 320mg/dl ethanol for 5 days prior to differentiation

treatment_protocol: Following treatment with 320mg/dl ethanol in mitogenic media, neurospheres were differentiated by the removal of EGF and LIF from the mitogenic media for 3 days without ethanol (treatment)

tag: Differentiated gestational Day 12.5 Mouse cortical-derived neurospheres pre-exposed to 320mg/dl ethanol (treatment)


title: Mouse neurospheres differentiated in the presence of FGF, b5 corresponds to replicate 7

description: Differentiated gestational Day 12.5 Mouse cortical-derived neurospheres


title: Mouse neurospheres differentiated in the presence of FGF, B3 corresponds to replicate 5


title: Mouse neurospheres differentiated in the presence of FGF, b1r corresponds to replicate 8


title: Mouse neurospheres differentiated in the presence of FGF, A1r corresponds to replicate 3


title: Mouse neurospheres differentiated in the presence of FGF, A3 corresponds to replicate 1

growth_protocol: Neurospheres were cultured in serum-free mitogenic medium DMEM/F-12, 20ng/ml basic Fibroblast Growth Factor, 20ng/ml human recombinant Epidermal Growth Factor, 0.15ng/ml human recombinant Leukemia Inhibitory Factor, 1X Insulin-Transferrin-Selenium-X Supplement, 5ug/ml heparin, and 20nM progesterone and cultured at 37C (5% CO2) in a humidified chamber to allow formation of neurospheres


title: Mouse neurospheres differentiated in the presence of FGF, A2r corresponds to replicate 4


title: Mouse neurospheres differentiated in the presence of FGF, A5 corresponds to replicate 2