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MB Sample ID: SA022994
Local Sample ID: | C_1_r6 |
Subject ID: | SU000475 |
Subject Type: | Human Cells |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Species Group: | Human |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU000475 |
Subject Type: | Human Cells |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Species Group: | Human |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
C_1_r6 | SA022994 | FL005598 | DMSO | Treatment |
C_1_r6 | SA022994 | FL005598 | Normal | Growth_type |
Collection:
Collection ID: | CO000469 |
Collection Summary: | An aliquot of the media following treatment was collected, and the remainder of the media was aspirated. The cells were washed with PBS twice, and quenched with 8mL ice-cold isotonic 0.9% (w/v) saline for 2 minutes. Total cellular content was then extracted with 1.7mL ice-cold acetonitrile/water (50:50, v/v) solution. Cell extracts were collected using a cell scraper and quickly transferred to MagNA Lyser Green Beads tubes (Roche, Indianapolis, USA) and stored in -80°C. Media was added to empty plates and incubated together with the cells for the duration of the experiment served as a blank. |
Sample Type: | Fibroblasts |
Treatment:
Treatment ID: | TR000489 |
Treatment Summary: | Monolayer cell cultures of desmoid tumors and normal fibroblasts from desmoid patients and an unaffected fibroblast cell line were grown in DMEM supplemented with 5% fetal bovine serum and maintained at 37°C in 5% CO2. Cells were divided when confluent and experiments were performed between the third and sixth passages. Approximately 10 x 106 cells were treated with 1.0uM Dasatinib (Selleck, Houston, USA) dissolved in DMSO, or 0.5uM FAK Inhibitor 14 (Cayman Chemicals Company, Michigan, USA) dissolved in water. Cells were incubated in fresh media containing the inhibitors, or vehicle, for 24 hours. |
Sample Preparation:
Sampleprep ID: | SP000482 |
Sampleprep Summary: | Cells were homogenized on the MagNA Lyzer, with two 30-sec cycles at 2000 rpm, resting in a -20 °C chilling block for 1 min in between pulses, and centrifuged samples at 16,000 rcf for 4 min. The cell lysate was transferred to a new 2 mL Lo-Bind Eppendorf tubes, with the final cell count approximately 10x10^6 cells for each sample. Of the twenty cell lysate samples, six samples had sufficient volume for study samples and to be included in an analytical quality control (QC) total pool. Aliquots from these cell lysate samples were combined, divided into three total pool aliquots, and processed identically to the cell lysate study samples. All study and pool samples were lyophilized to dryness and reconstituted in a 0.2M phosphate buffer, pH 7.4, in D2O with 10% Chenomx ISTD. |
Analysis:
MB Sample ID: | SA022994 |
Analysis ID: | AN000711 |
Analysis Type: | NMR |
Num Factors: | 13 |
NMR:
NMR ID: | NM000078 |
Analysis ID: | AN000711 |
Instrument Name: | Bruker Avance III |
Instrument Type: | FT-NMR |
NMR Experiment Type: | 1D 1H |
Spectrometer Frequency: | 700 MHz |