NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM21897 Query DataSets for GSM21897
Status Public on Apr 29, 2004
Title Matrigel induced differentiation of HBC-3 cells , time 5-A days
Sample type RNA
 
Channel 1
Source name HBC-3 cells after 5 day of differentiation
Organism Mus musculus
Extracted molecule total RNA
 
Channel 2
Source name HBC-3 cells control
Organism Mus musculus
Extracted molecule total RNA
 
 
Description HBC-3 cells were maintained as previously described (Rogler,L.E.,Am.J. Path. 150:591-602). Cells grown for microarray analysis were stripped of their STON+ feeder layer by trypsinization and replating on plastic in HBM
Undifferentiated cells were harvested the next A.M. for preparation of total RNA. Twenty 10cm plates were used to obtain sufficient quantities of undifferentiated reference total RNA. Total RNAs were prepared using Qiagen RNeasy maxi kit. The quality of each RNA preparation was determined using an Agilent 2100 Bioanalyzer. Five microgram aliquots of total RNA from undifferentiated HBC-3 cells were used to generate amplified RNA (aRNA) using the Ambion Message Amp Kit (cat.# 1750). The quality of the aRNA was determined using an Agilent Bioanalyzer. Amplified RNAs were stored as 5ug aliquots under 100% Ethanol at -80oC.
Cy 3 and Cy5 labeled targets were synthesized using a modified version of the AECOM functional Genomics facility protocol (http://microarray1k.aecom.yu.edu/) as previously described (Plescia et al, 2001,Differentiation 68:254-269). Cy3 and Cy5 labeled targets were prepared using 5 ug of aRNA using random primers and Superscript reverse transcriptase (Invitrogen)
Cy3 labeled target prepared from undifferentiated HBC-3 cells provided the reference sample for all comparisons. The precipitated targets were then resuspended in hybridization buffer as previously described and applied to a microarray. Arrays were hybridized at 50oC for 20 hours. Washing was carried out as described at (http://microarray1k.aecom.yu.edu/
Cy3 and Cy5 labeled targets were synthesized using a modified version of the AECOM functional Genomics facility protocol (http://microarray1k.aecom.yu.edu/) as previously described (Plescia et al, 2001,Differentiation 68:254-269). Cy3 and Cy5 labeled targets were prepared using 5 ug of aRNA using random primers and Superscript reverse transcriptase (Invitrogen).). The purified Cy5 labeled targets were repared from aRNA made from HBC-3 cells treated with Matrigel for 5 day (to induce bile duct differentiation).Cy3 reference target and a Cy5 target prepared from an RNA isolated from cells at one of the time points of Matrigel treatment were combined and precipitated. The precipitated targets were then resuspended in hybridization buffer as previously described and applied to a microarray. Arrays were hybridized at 50oC for 20 hours. Washing was carried out as described at (http://microarray1k.aecom.yu.edu/). "
In vitro bile ductular differentiation of HBC-3 cells was performed as previously described (Rogler,L.E.,Am.J. Path. 150:591-602) Cells grown for microarray analysis were stripped of their STON+ feeder layer by trypsinization and on a 1mm Matrigel cushion (differentiated). Cells undergoing Matrigel induced differentiation were harvested at 1 day of Matrigel treatment for preparation of total RNA. Ten 10 cm plates were harvested for each time point to obtain total RNA sufficient for several target preparations at each time point. Five microgram aliquots of total RNA from undifferentiated HBC-3 or Matrigel treated HBC-3 cells were used to generate amplified RNA (aRNA) using the Ambion Message Amp Kit (cat.# 1750). The quality of the aRNA was determined using an Agilent Bioanalyzer. Amplified RNAs were stored as 5ug aliquots under 100% Ethanol at -80oC.
Following hybridization and washing, the arrays were scanned for Cy3 and y5 florescence using a scanner Genepix 3.0 scanner (Axon Instruments Inc., Union City, CA). The signal intensity of a spot was considered to be significant if the signal intensity was greater by 200 units of the background for that spot and if it was not flagged as ""not found"" by GenePix software. These data were normalized using the Lowess implementation included in BASE. Following filtration and normalization, the ratio of red and green fluorescence, (RAT) was quantified for each qualified gene. The RAT values were then Log transformed and the LOG2 of the RAT is presented in the data table."
Keywords = HBC-3 cells, murine hepatoblast cells, matrigel
 
Submission date Apr 27, 2004
Last update date May 27, 2005
Contact name Leslie Rogler
E-mail(s) rogler@aecom.yu.edu
Phone 718 430 3651
Organization name Albert Einstein College of Medicine
Department Liver Research Center/ Department of Medicine
Street address 1300 Morris Park Ave
City New York
State/province NY
ZIP/Postal code 10461
Country USA
 
Platform ID GPL1205
Series (1)
GSE1348 HBC-3 matrigel differentiation

Data table header descriptions
ID_REF
VALUE Normalized log2 ratio of medians after background substraction
dia spot diameter
FCh1Median Channel 1 median intensity
BCh1Median Channel 1 median background intensity
FCh2Median Channel 2 median intensity
BCh2Median Channel 2 median background intensity

Data table
ID_REF VALUE dia FCh1Median BCh1Median FCh2Median BCh2Median
1 -0.33665984 110 327 40 373 42
2 0.71756631 110 930 41 512 43
3 1.72221725 110 714 42 244 42
4 0.60225776 110 1418 42 841 42
5 0.06082972 120 3321 42 2871 42
7 0.1362835 110 2456 41 2090 41
11 0.46112387 110 744 41 497 41
12 -0.01933101 100 317 40 284 40
14 -0.05286912 110 4218 43 3824 43
16 0.09772585 120 12660 49 11449 50
42 0.2146323 120 15086 48 13408 46
23936 -0.24015152 140 12759 48 16289 49
17 -0.08873604 120 809 48 783 50
18 0.05357703 120 11388 64 11012 63
19 -0.5042085 100 1693 43 2193 44
20 0.33825944 110 2648 50 1943 46
21 -0.0653426 110 747 43 697 42
22 -0.93921714 120 1379 41 2280 41
23 1.50128452 160 1631 42 559 41
30 -0.30764053 100 2580 45 3059 45

Total number of rows: 17845

Table truncated, full table size 631 Kbytes.




Supplementary data files not provided

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap