Search results for the GEO ID: GSE13738  |
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GSM ID | GPL ID |
Select for analysis |
Title |
Source name |
Description |
Characteristics |
GSM345232 | GPL570 |
|
T cell directly activated_rep1
|
T cell directly activated
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345232
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345232/suppl/GSM345232.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345246 | GPL570 |
|
T cell bystander activated_rep2
|
T cell bystander activated
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345246
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345246/suppl/GSM345246.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345250 | GPL570 |
|
T cell bystander activated_rep3
|
T cell bystander activated
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345250
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345250/suppl/GSM345250.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345259 | GPL570 |
|
T cell directly activated_rep2
|
T cell directly activated
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345259
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345259/suppl/GSM345259.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345264 | GPL570 |
|
T cell bystander activated_rep1
|
T cell bystander activated
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345264
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345264/suppl/GSM345264.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345266 | GPL570 |
|
T cell directly activated_rep3
|
T cell directly activated
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345266
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345266/suppl/GSM345266.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345267 | GPL570 |
|
T cell bystander activated_rep4
|
T cell bystander activated
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345267
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345267/suppl/GSM345267.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345280 | GPL570 |
|
T cell directly activated_rep4
|
T cell directly activated
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345280
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345280/suppl/GSM345280.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345281 | GPL570 |
|
Resting T cells_rep1
|
Resting cells
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
Resting T cells - cells expressing a TCR which does not specifically recognize SEB (Vβ13.1 T cells) and did not upregulate the activation marker CD25 in response to transwell SEB-stimulated co-culture over a 5 day period.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345281
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345281/suppl/GSM345281.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345282 | GPL570 |
|
Resting T cells_rep2
|
Resting T cells
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads.
Resting T cells - cells expressing a TCR which does not specifically recognize SEB (Vβ13.1 T cells) and did not upregulate the activation marker CD25 in response to transwell SEB-stimulated co-culture over a 5 day period.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345282
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345282/suppl/GSM345282.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345283 | GPL570 |
|
Resting T cells_rep3
|
Resting T cells
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads. Resting T cells - cells expressing a TCR which does not specifically recognize SEB (Vβ13.1 T cells) and did not upregulate the activation marker CD25 in response to transwell SEB-stimulated co-culture over a 5 day period.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345283
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345283/suppl/GSM345283.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
GSM345284 | GPL570 |
|
Resting T cells_rep4
|
Resting T cells
|
Briefly, PBMCs were isolated from buffy coats by density gradient centrifugation. Vβ13.1 and Vβ17 expressing T cells were enriched using PE-conjugated antibodies with MACS anti-PE microbeads. Resting T cells - cells expressing a TCR which does not specifically recognize SEB (Vβ13.1 T cells) and did not upregulate the activation marker CD25 in response to transwell SEB-stimulated co-culture over a 5 day period.
|
we set up an in vitro system within which to analyse bystander T cell activation in human T cells, in the absence of the possibility for TCR cross-reactivity. In addition, we have investigated the genetic, phenotypic, and functional characteristics of bystander activated T cells.
|
| Sample_geo_accession | GSM345284
| | Sample_status | Public on Feb 11 2009
| | Sample_submission_date | Nov 25 2008
| | Sample_last_update_date | Feb 11 2009
| | Sample_type | RNA
| | Sample_channel_count | 1
| | Sample_organism_ch1 | Homo sapiens
| | Sample_taxid_ch1 | 9606
| | Sample_biomaterial_provider_ch1 | UK National Blood Centre
| | Sample_treatment_protocol_ch1 | Vβ13.1+ T cells were plated out in the top well of transwell inserts and co-cultured with whole autologous PBMC plated in the lower well. Vβ17+ T cells were plated out in 24 well plates mixed with whole autologous PBMC, present at an identical ratio of PBMC to purified T cells to that used in the transwell culture. All wells were treated with 2.5 μg/ml SEB. Cultures were incubated for five days at 37 ˚C in the presence of 5% CO2, then the top wells of transwell cultures and whole mixed wells were harvested and stained. Cells were sorted by FACS into bystander activated “B” (CD3+, Vβ13.1+, CD25+), resting “R” (CD3+, Vβ13.1+, CD25-) and directly activated “D” (CD3+, Vβ17+, CD25+) populations.
| | Sample_growth_protocol_ch1 | T cells expressing Vβ17 TCRs respond specifically to stimulation with the superantigen staphylococcal enterotoxin B (SEB) – this method was used to generate the “directly activated” sample of T cells. T cells expressing Vβ13.1 TCRs do not respond to direct SEB stimulation, however, when cultured with soluble factors produced by whole PBMC stimulated with SEB, a proportion of Vβ13.1 T cells will upregulate activation markers – these cells are used as the “bystander activated” T cell sample.The “Resting” T cell sample consists of those Vβ13.1 T cells which do not upregulate activation markers in response to the bystander stimulus.
| | Sample_molecule_ch1 | total RNA
| | Sample_extract_protocol_ch1 | RNA was extracted using RNeasy kits (Qiagen).
| | Sample_label_ch1 | Biotin
| | Sample_label_protocol_ch1 | Two-cycle cDNA synthesis kit (Affymetrix # 900494) used to convert 50ng total RNA together with appropriate dilutions of GeneChip eukaryotic poly-A RNA spike controls (Affymetrix # 900433) to double-stranded cDNA with superscript II using T7-(dT)24 primer. This was used in an in vitro transcription to generate biotin-labelled cRNA probes.
| | Sample_hyb_protocol | Fragmented labelled cRNA (15µg) was used in 300µl hybridization cocktail containing spiked controls (Affymetrix #900299). A 200µl of this hybridization cocktail was used on the chip and incubated at 45°C for 16 h in the hybridization oven, rotating at 60 rpm. Following hybridization, the arrays were processed using a Genechip Fluidics Station 400 according to recommended protocols (EukGE-WS2v5, Affymetrix) of double-staining and post-hybridisations washes.
| | Sample_scan_protocol | Fluorescent images were captured using gene Array Scanner 3000 and GCOS1.2 software (Affymetrix).
| | Sample_data_processing | We have used the genechip robust multi-array average expression measurements (RMA) as implemented in BioConductor R statistics (www.bioconductor.org). Cell Intensity files (CEL) were used to obtain expression values for all the 54,675 probe sets (transcripts) on each of these arrays.
| | Sample_platform_id | GPL570
| | Sample_contact_name | Dilair,,Baban
| | Sample_contact_email | dilair.baban@well.ox.ac.uk
| | Sample_contact_phone | +44(0)1865287521
| | Sample_contact_fax | +44(0)1865287501
| | Sample_contact_laboratory | Genomics
| | Sample_contact_department | Wellcome Trust Centre Human Genetics
| | Sample_contact_institute | University of Oxford
| | Sample_contact_address | Roosevelt Drive
| | Sample_contact_city | Oxford
| | Sample_contact_zip/postal_code | OX3 7BN
| | Sample_contact_country | United Kingdom
| | Sample_supplementary_file | ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM345nnn/GSM345284/suppl/GSM345284.CEL.gz
| | Sample_series_id | GSE13738
| | Sample_data_row_count | 54675
| | |
|
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