|
| Status |
Public on Dec 31, 2014 |
| Title |
CD24-low/CD44-high (CD44H) cell fraction 3 |
| Sample type |
RNA |
| |
|
| Source name |
CD24-low/CD44-high (CD44H) cell fraction
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: esophagus
cell type: transformed epithelial cells (keratinocytes)
cell fraction: CD44H
genotype: EGFR-cyclin D1-p53R175H-hTERT-DNMAML1
|
| Treatment protocol |
To analyze CD24 and CD44 expression, cells were detached by Versene (0.02% Tetrasodium ethylenediaminetetraacetate in Phosphate Buffered Saline) and incubated with Allophycocyanin-conjugated anti-human CD44 (BD Bioscience) and PE/Cy7-conjugated anti human CD24 (Biolegend, San Diego, CA) on ice for 30 min. The cells were washed with Hank's Buffered Salt Solution (HBSS) containing 1% bovine serum albumin (BSA) twice and then detected by FACSCalibur (BD Bioscience). The CD24/CD44 stained cells were sorted at the Flow cytometry facility of the University of Pennsylvania.
|
| Growth protocol |
All cells were grown under regular conditions (21% O2, 5% CO2, 95% humidity) at 37°C in serum free medium (Keratinocyte-SFM)(Invitrogen) supplemented with 1 ng/ml epidermal growth factor and 50 µg/ml bovine pituitary extract.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNeasy Mini Kit (Qiagen, Inc., Valencia, CA, USA) was used to extract total RNA according to the manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol.
|
| |
|
| Hybridization protocol |
Hybridization of cRNAs to Affymetrix chips were performed according manufacturer's instructions
|
| Scan protocol |
Scanning of Affymetrix chips were performed according manufacturer's instructions
|
| Description |
4801CD44L3
|
| Data processing |
.CEL files (probe level data) were exported from Affymetrix® GeneChip® Operating Software which was also used to calculate detection flags, which were exported in .chp files. Probe level data (.CEL files) were imported with Partek Genomics Suite (v. 6.4, Partek, Inc., St. Louis, MO). The data were normalized and summarized to the probe set level using GC-RMA, yielding log2 intensities for all probe sets on the array.
|
| |
|
| Submission date |
Apr 22, 2011 |
| Last update date |
Dec 31, 2014 |
| Contact name |
Hiroshi Nakagawa |
| E-mail(s) |
nakagawh@mail.med.upenn.edu
|
| Phone |
215-573-1867
|
| Organization name |
University of Pennsylvania
|
| Department |
Gastroenterology Division
|
| Lab |
Nakagawa
|
| Street address |
415 Curie Blvd.
|
| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19104 |
| Country |
USA |
| |
|
| Platform ID |
GPL6244 |
| Series (1) |
| GSE28802 |
CD24-low Cancer stem-cell like cell fraction in Notch inhibited transformed human esophageal cells |
|
