|
Status |
Public on Jan 01, 2021 |
Title |
thigh; stromal pellet; F83 |
Sample type |
RNA |
|
|
Channel 1 |
Source name |
Stratagene Human Universal Reference, 10 ug labeled aRNA
|
Organism |
Homo sapiens |
Characteristics |
reference: Stratagene Human Universal Reference
|
Extracted molecule |
total RNA |
Extraction protocol |
not provided
|
Label |
Cy3
|
Label protocol |
not provided
|
|
|
Channel 2 |
Source name |
thigh; stromal pellet; F83, 10 ug labeled aRNA
|
Organism |
Homo sapiens |
Characteristics |
age: 83 years organism part: thigh sample fraction: stromal pellet Sex: female
|
Extracted molecule |
total RNA |
Extraction protocol |
Adipose Tissue Samples; Specimen Collection Adipose tissue specimens were collected either from the Stanford Tissue Bank, Stanford Dermatology Clinic, Stanford Surgical Pathology or directly from the surgeon at the time of surgery. All surgeries were performed at Stanford Hospital and Clinics. When necessary a trained pathologist dissected the desired adipose specimen from the organ/limb/skin to which it was attached. Samples acquired by Stanford Tissue Bank or Surgical Pathology may have been stored at 4C for up to several hours but were always processed on the same day as the surgery was performed. All samples are considered normal human tissues and were originally collected under the IRB Study of Messenger RNA Expression in Normal Human Tissues ID: 76363, which did not require patient consent as very little patient information (age, gender, procedure) was provided. Subsequently it was determined that our sample collection does not meet the definitions of human subject research under Stanfords Human Research Protection Program and therefore an IRB protocol was no longer required. Tissue Fractionation - Clemons; Tissue Fractionation Adipose tissue samples were thoroughly rinsed with 70% ethanol to minimize risk of microbial contamination of cell cultures. Tissues were then rinsed with Hanks Balanced Salt Solution (HBSS) and transferred to a clean dish. At this time 1-4 g of tissue were removed and stored at -80C. Additionally a thin slice of each tissue was placed in 10% formalin for paraffin embedding. The remainder of each tissue sample was minced using surgical scissors. Tissue pieces were transferred to 50 ml conical tubes (up to 25 ml tissue per tube), and an equivalent volume of collagenase solution* was added to the tube. Tissue was again minced with scissors until no large pieces remained. Tubes were shaken to mix and incubated at 37C for 30 min to 4 hours, until pieces of tissue were no longer visible. Shin/calf adipose tissue in general took the longest and abdominal and thigh lipoaspirate the shortest time to digest. Digested tissues were then passed through a 180-micron nylon net filter and centrifuged at 100g for 5 minutes to pellet stromal-vascular cells. A 16-gauge needle attached to a 20 ml syringe was passed through the top visible lipid/adipocyte layer and the entire bottom layer containing buffer, cell pellet and blood cells was aspirated from the bottom and transferred to a new conical, tube. This cell suspension (stromal-vascular fraction) was thoroughly mixed by pipetting and divided equally into 2 conical tubes. One of these was set aside for cell culture (see Cell Culture) and the other was centrifuged at 400g for 7 minutes. Buffer was carefully aspirated with a pipette to avoid losing pelleted cells and 15 ml Trizol was added. The remaining lipid/adipocyte layer was washed 1-2 times depending on the amount of visible blood cells: 25 ml HBSS was added to the adipocytes, and gently inverted to mix. Adipocytes were centrifuged at 100g and excess buffer along with contaminating cells was aspirated from the bottom using a needle and syringe as described above. An equal volume of Trizol was added to the washed adipocytes. The stromal-vascular pellet sample and the adipocyte sample were stored in Trizol at -80C. Collagenase Solution: For stock solution 1000 U/ml Sigma Type II Collagenase (Cat# C6885) was dissolved in HBSS and centrifuged at 1000g for 20 minutes. Resulting liquid fraction was filtered (.22 micron) and stored at -20C and pelleted crystals were discarded. For working solution 1000 U/ml collagenase stock solution was diluted 3x in HBSS containing 5mM CaCl2 Amplification 1 ug of each total RNA sample was amplified using Ambion Amino Allyl MessageAmp II-96 Kit (Cat# 1821) according to manufacturers protocol
|
Label |
Cy5
|
Label protocol |
not provided
|
|
|
|
Hybridization protocol |
not provided
|
Scan protocol |
Model: GenePix 4000B
|
Description |
Simple annotation: Normal tissue, fat image: http://smd.stanford.edu/MicroArray/gifs/2008-04/79731.gif
|
Data processing |
VALUE is Log (base 2) of the ratio of the median of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm)
|
|
|
Submission date |
Nov 08, 2010 |
Last update date |
Jan 01, 2021 |
Contact name |
Heather Clemons |
E-mail(s) |
hjeanc@gmail.com
|
Phone |
(650) 723-0005
|
Organization name |
Stanford School of Medicine
|
Department |
Dept of Biochemistry
|
Street address |
Beckman Center
|
City |
Stanford |
State/province |
CA |
ZIP/Postal code |
94305 |
Country |
USA |
|
|
Platform ID |
GPL11179 |
Series (1) |
GSE25194 |
Expression profiles of human adipose tissue, adipocytes and stromal-vascular pellet cells from multiple body sites |
|
Data table header descriptions |
ID_REF |
ID_REF |
CH1I_MEAN |
Mean feature pixel intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale |
CH2I_MEAN |
Mean feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale |
CH1B_MEDIAN |
The median feature background intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel; Background |
CH2B_MEDIAN |
The median feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel; Background |
CH1D_MEAN |
The mean feature pixel intensity at wavelength 532 nm with the median background subtracted.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel |
CH2D_MEAN |
.The mean feature pixel intensity at wavelength 635 nm with the median background subtracted.; Type: integer; Scale: linear_scale; Channel: Cy5 channel |
CH1I_MEDIAN |
Median feature pixel intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale |
CH2I_MEDIAN |
Median feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale |
CH1B_MEAN |
The mean feature background intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Background |
CH2B_MEAN |
The mean feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Background |
CH1D_MEDIAN |
The median feature pixel intensity at wavelength 532 nm with the median background subtracted.; Type: integer; Scale: linear_scale |
CH2D_MEDIAN |
The median feature pixel intensity at wavelength 635 nm with the median background subtracted.; Type: integer; Scale: linear_scale |
CH1_PER_SAT |
The percentage of feature pixels at wavelength 532 nm that are saturated.; Type: integer; Scale: linear_scale |
CH2_PER_SAT |
The percentage of feature pixels at wavelength 635 nm that are saturated.; Type: integer; Scale: linear_scale |
CH1I_SD |
The standard deviation of the feature intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel |
CH2I_SD |
The standard deviation of the feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel |
CH1B_SD |
The standard deviation of the feature background intensity at wavelength 532 nm.; Type: float; Scale: linear_scale; Channel: Cy3 Channel; Background |
CH2B_SD |
The standard deviation of the feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel; Background |
PERGTBCH1I_1SD |
The percentage of feature pixels with intensities more than one standard deviation above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale |
PERGTBCH2I_1SD |
The percentage of feature pixels with intensities more than one standard deviation above the background pixel intensity, at wavelength 635 nm.; Type: integer; Scale: linear_scale |
PERGTBCH1I_2SD |
The percentage of feature pixels with intensities more than two standard deviations above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale |
PERGTBCH2I_2SD |
The percentage of feature pixels with intensities more than two standard deviations above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale |
SUM_MEAN |
The sum of the arithmetic mean intensities for each wavelength, with the median background subtracted.; Type: integer; Scale: linear_scale |
SUM_MEDIAN |
The sum of the median intensities for each wavelength, with the median background subtracted.; Type: integer; Scale: linear_scale |
RAT1_MEAN |
Ratio of the arithmetic mean intensities of each spot for each wavelength, with the median background subtracted. Channel 1/Channel 2 ratio, (CH1I_MEAN - CH1B_MEDIAN)/(CH2I_MEAN - CH2B_MEDIAN) or Green/Red ratio.; Type: float; Scale: linear_scale |
RAT2_MEAN |
The ratio of the arithmetic mean intensities of each feature for each wavelength, with the median background subtracted.; Type: float; Scale: linear_scale |
RAT2_MEDIAN |
The ratio of the median intensities of each feature for each wavelength, with the median background subtracted.; Type: float; Scale: linear_scale |
PIX_RAT2_MEAN |
The geometric mean of the pixel-by-pixel ratios of pixel intensities, with the median background subtracted.; Type: float; Scale: linear_scale |
PIX_RAT2_MEDIAN |
The median of pixel-by-pixel ratios of pixel intensities, with the median background subtracted.; Type: float; Scale: linear_scale |
RAT2_SD |
The geometric standard deviation of the pixel intensity ratios.; Type: float; Scale: linear_scale |
TOT_SPIX |
The total number of feature pixels.; Type: integer; Scale: linear_scale |
TOT_BPIX |
The total number of background pixels.; Type: integer; Scale: linear_scale |
REGR |
The regression ratio of every pixel in a 2-feature-diameter circle around the center of the feature.; Type: float; Scale: linear_scale |
CORR |
The correlation between channel1 (Cy3) & Channel 2 (Cy5) pixels within the spot, and is a useful quality control parameter. Generally, high values imply better fit & good spot quality.; Type: float; Scale: linear_scale |
DIAMETER |
The diameter in um of the feature-indicator.; Type: integer; Scale: linear_scale |
X_COORD |
X-coordinate of the center of the spot-indicator associated with the spot, where (0,0) is the top left of the image.; Type: integer; Scale: linear_scale |
Y_COORD |
Y-coordinate of the center of the spot-indicator associated with the spot, where (0,0) is the top left of the image.; Type: integer; Scale: linear_scale |
TOP |
Box top: int(((centerX - radius) - Xoffset) / pixelSize).; Type: integer; Scale: linear_scale |
BOT |
Box bottom: int(((centerX + radius) - Xoffset) / pixelSize).; Type: integer; Scale: linear_scale |
LEFT |
Box left: int(((centerY - radius) - yoffset) / pixelSize).; Type: integer; Scale: linear_scale |
RIGHT |
Box right: int(((centerY + radius) - yoffset) / pixelSize); Type: integer; Scale: linear_scale |
FLAG |
The type of flag associated with a feature: -100 = user-flagged null spot; -50 = software-flagged null spot; 0 = spot valid.; Type: integer; Scale: linear_scale |
CH2IN_MEAN |
Normalized value of mean Channel 2 (usually 635 nm) intensity (CH2I_MEAN/Normalization factor).; Type: integer; Scale: linear_scale; Channel: Cy5 channel |
CH2BN_MEDIAN |
Normalized value of median Channel 2 (usually 635 nm) background (CH2B_MEDIAN/Normalization factor).; Type: integer; Scale: linear_scale; Channel: Cy5 channel; Background |
CH2DN_MEAN |
Normalized value of mean Channel 2 (usually 635 nm) intensity with normalized background subtracted (CH2IN_MEAN - CH2BN_MEDIAN).; Type: integer; Scale: linear_scale; Channel: Cy5 channel |
RAT2N_MEAN |
Type: float; Scale: linear_scale |
CH2IN_MEDIAN |
Normalized value of median Channel 2 (usually 635 nm) intensity (CH2I_MEDIAN/Normalization factor).; Type: integer; Scale: linear_scale |
CH2DN_MEDIAN |
Normalized value of median Channel 2 (usually 635 nm) intensity with normalized background subtracted (CH2IN_MEDIAN - CH2BN_MEDIAN).; Type: integer; Scale: linear_scale |
RAT1N_MEAN |
Ratio of the means of Channel 1 (usually 532 nm) intensity to normalized Channel 2 (usually 635 nm) intensity with median background subtracted (CH1D_MEAN/CH2DN_MEAN). Channel 1/Channel 2 ratio normalized or Green/Red ratio normalized.; Type: float; Scale: linear_scale |
RAT2N_MEDIAN |
Channel 2/Channel 1 ratio normalized, RAT2_MEDIAN/Normalization factor or Red/Green median ratio normalized.; Type: float; Scale: linear_scale |
LOG_RAT2N_MEAN |
Log (base 2) of the ratio of the mean of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm) [log (base 2) (RAT2N_MEAN)].; Type: float; Scale: log_base_2 |
VALUE |
Log (base 2) of the ratio of the median of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm) [log (base 2) (RAT2N_MEDIAN)].; Type: float; Scale: log_base_2 |