|
| Status |
Public on Dec 31, 2008 |
| Title |
naïve B cells, rep 1 |
| Sample type |
RNA |
| |
|
| Source name |
sort-purified human splenic naïve B cells
|
| Organism |
Homo sapiens |
| Characteristics |
Untreated
Tissue: human spleen
|
| Treatment protocol |
Naïve and memory B cell subsets were isolated by cell sorting using antibodies to CD20, CD27 and Ig isotypes. Cells were washed in PBS.
|
| Growth protocol |
Human splenic tissue (Australian Red Cross) was used as a source of mature immune cells. Tissue was dissected and capsule disrupted. Red blood cells were lysed and cells washed in RPMI-10%FCS. MNCs were isolated by collecting the buffy coat after layering on Ficoll-Paque.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were lysed and RNA extracted using the RNeasy mini kit (Qiagen) according to the manufacturer's instructions, with the inlcusion of the DNase step.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to a modified protocol published (Baugh et al, 2001) using an in vitro transcription kit (Enzo Biosciences) that amplifies and labels cRNA from 1 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).
|
| |
|
| Hybridization protocol |
Following fragmentation, 20 ug of cRNA were hybridized for 16 hr at 45C on Human Genome U133 set GeneChips. GeneChips were washed and stained in the Affymetrix GeneChip Fluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using the GeneChip Scanner 3000 7G.
|
| Description |
Replicate 1 of 2
|
| Data processing |
The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method, following which absolute expression or comparison analysis were performed by the software using its statistical expression algorithm. Genes with a fold change of 2 or more, p<0.05, and a present signal of more than 100 were examined.
|
| |
|
| Submission date |
Oct 30, 2008 |
| Last update date |
Sep 01, 2016 |
| Contact name |
Stuart Tangye |
| E-mail(s) |
s.tangye@garvan.org.au
|
| Phone |
+61 2 9295 8455
|
| Organization name |
Garvan Institute of Medical Research
|
| Department |
Immunology
|
| Street address |
384 Victoria St
|
| City |
Darlinghurst |
| State/province |
NSW |
| ZIP/Postal code |
2010 |
| Country |
Australia |
| |
|
| Platform ID |
GPL96 |
| Series (1) |
| GSE13411 |
Gene expression by human splenic B-cell subsets |
|
| Relations |
| Reanalyzed by |
GSE86363 |
