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Sample GSM4214045 Query DataSets for GSM4214045
Status Public on Dec 12, 2019
Title EyeBank_cultured_human_corneal_epithelial_cells_006
Sample type RNA
Source name primary human epithelial corneal cell population
Organism Homo sapiens
Characteristics age: 71
Sex: female
protocol: cultivated 14 days without feeder layer
Treatment protocol Total RNA was isolated using the RNeasy Mini Kit (QIAGEN, Cat#74104)
Growth protocol Once the Descement membrane has been peeled, remaining corneas (stromas with their attached epithelial cells) are then incubated with dispase (2.5 mg/ml) overnight at 4°C. Using forceps, the epithelium is mechanically separated from the underlying stroma and treated with trypsin, which allows the human corneal epithelial cells (hCECs) to dissociate one from another. Once isolated, hCECs are seeded in tissues culture flasks together with feeder layer for expansion.
Cells are grown in DH medium (Dulbecco–Vogt modification of Eagle's medium with Ham's F12 in a 3:1 ratio supplemented with 5% FetalClone II serum, 5 μg/ml of insulin, 0.4 μg/ml of hydrocortisone, 10 ng/ml of epidermal growth factor, 10−10 M of cholera toxin, 100 IU/ml of penicillin, and 25 μg/ml of gentamycin) until they reach 75-95% confluence.
Extracted molecule total RNA
Extraction protocol Cyanine-3 (Cy3) labeled cRNA was prepared from 50 ng RNA using the One-Color LowInput QuickAmp Labeling Kit One-Color (Agilent) according to the manufacturer's instructions
Label Cy3
Label protocol 600 ng of Cy3-labelled cRNA (specific activity >8.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 25 ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole SurePrint G3 Human GE 8x60K (G4851A) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately.
Hybridization protocol Slides were scanned immediately after washing on the Agilent SureScanner (G4900DA) using one color scan setting for 1x60k array slides (Scan resolution 3um, Dye channel is set to Green and Green PMT is set to 100%).
Scan protocol The scanned images were analyzed with Feature Extraction Software 10,7 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
Description Gene expression of passage 3 cells cultivated 14 days without feeder layer (80% confluence)
Data processing gPixNormIQR : The normalized Inter-quartile range of all of the inlier pixels per feature. The range is computed independently in each channel.
Submission date Dec 11, 2019
Last update date Dec 12, 2019
Contact name Gaëtan Le-Bel
Organization name CUO-Recherche
Lab Sylvain Guérin
Street address Local H2-00, Hôpital du Saint-Sacrement,1050 Chemin Sainte-Foy
City Québec
State/province Quebec
ZIP/Postal code G1S 4L8
Country Canada
Platform ID GPL13607
Series (1)
GSE141841 Gene expression of human corneal epithelial cells grown with irradiated human fibroblasts or irradiated mouse 3T3 feeder layer .

Data table header descriptions
VALUE normalized signal

Data table
1 6051.23200
2 17.42055
3 13.89937
4 51.14970
5 14.45535
6 21.49770
7 145.29480
8 377.32170
9 12.97275
10 19.64445
11 17.42055
12 110.45370
13 154.19040
14 144.36820
15 74.13000
16 17.42055
17 24.46290
18 11.86080
19 21.49770
20 44.84865

Total number of rows: 62976

Table truncated, full table size 924 Kbytes.

Supplementary file Size Download File type/resource
GSM4214045_SG11400001_252800422108_S001_GE1_107_Sep09_2_4.txt.gz 12.3 Mb (ftp)(http) TXT
Processed data included within Sample table

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