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Status |
Public on Sep 27, 2013 |
Title |
CPE-human-7 |
Sample type |
RNA |
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Source name |
healthy human choroid plexus
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Organism |
Homo sapiens |
Characteristics |
tissue: choroid plexus epithelium cell sampling: laser dissection microscopy
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Treatment protocol |
With laser dissection microscopy, the CPE cells were specifically cut out from cryosections of the choroid plexus (10um) mounted on PEN-membrane slides
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Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was isolated using RNeasy Micro Kit (Qiagen Benelux, Venlo, The Netherlands). Subsequently, the mRNA component was amplified with Amino Allyl MessageAmp II aRNA Amplification kit (#1753, Ambion Applied Biosystems Europe B.V.).
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Label |
Cy5
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Label protocol |
aRNA samples were labeled with a Cy5 fluorescent probe (Cy5 mono-reactive dye pack, GE Healthcare UK, Little Chalfont, Buckinghamshire, UK). In short, aRNA was coupled to Cy5 (dissolved in DMSO) in 0.5 M NaHCO3 (pH 9.0) for one hour. Free dyes were quenched by the addition of 4 M hydroxylamine and removed by filtration over a Chroma Spin-30 column (DEPC-H20 Columns, Clontech Laboratories) with glycogen as a carrier.
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Hybridization protocol |
Labeled human CPE RNA and labeled human common reference sample were applied to microarrays enclosed in Agilent SureHyb-enabled hybridization chambers. After hybridization, slides were washed sequential
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Scan protocol |
Scanned on an Agilent scanner, protocol GE2-v5_95_Fec07
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Description |
reference ch2: tissue: RPE/chodoid, Cy3 CPE replicate 7 of 7 of healthy human choroid plexus
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Data processing |
Agilent Feature Extraction Software (v 9.5.3.1) was used for background subtraction and linear loweless dye normalization. This resulted amongst others in normalized log ratios (test/reference) data that can be find in the Agilent Feature Extraction files. Further data processing was done in R (version 2.14.0 for Windows, R Development Core Team, 2009), with package Limma. We read in the raw data (RG <-read.maimage). Next, we normalized within arrays (method 'loess', resulating in MA) and between arrays (method aquantile). In the matrix table, we listed these normalized red channels of each human CPE sample (log2).
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Submission date |
Aug 19, 2013 |
Last update date |
Sep 27, 2013 |
Contact name |
Sarah Janssen |
E-mail(s) |
s.janssen@nin.knaw.nl
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Organization name |
Netherlands Institute for Neuroscience
|
Department |
Ophthalmogenetics
|
Street address |
Meibergdreef 47
|
City |
Amsterdam |
ZIP/Postal code |
1105 BA |
Country |
Netherlands |
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Platform ID |
GPL4133 |
Series (2) |
GSE49969 |
Gene expression of the human choroid plexus epithelium (CPE) |
GSE49974 |
Gene expression of the choroid plexus epithelium (CPE) |
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