Return to study ST000442 main page
MB Sample ID: SA022544
| Local Sample ID: | TB468CDM48R1N |
| Subject ID: | SU000463 |
| Subject Type: | Cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Cell Biosource Or Supplier: | American Type Culture Collection |
| Cell Strain Details: | Breast Cancer Cell lines (BT474, MCF-7, MDA-MB-231, MDA-MB-468) |
| Cell Primary Immortalized: | Immortalized |
| Species Group: | Human |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU000463 |
| Subject Type: | Cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Cell Biosource Or Supplier: | American Type Culture Collection |
| Cell Strain Details: | Breast Cancer Cell lines (BT474, MCF-7, MDA-MB-231, MDA-MB-468) |
| Cell Primary Immortalized: | Immortalized |
| Species Group: | Human |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| TB468CDM48R1N | SA022544 | FL006431 | Media alone | Treatment |
Collection:
| Collection ID: | CO000457 |
| Collection Summary: | Cell samples were scraped off the dish in 4 mL of 50:50 acetonitrile:water and collected into 15 mL tubes prior to homogenization and extraction. |
| Sample Type: | cells |
| Storage Conditions: | -80°C |
Treatment:
| Treatment ID: | TR000477 |
| Treatment Summary: | Briefly, mycoplasma tested and certified BCa cell lines will be purchased from American Type Culture Collection (ATCC) and maintained under the manufacturer’s recommended conditions. Cells were grown for 48 h in the presence or absence of chemotherapeutic agent paclitaxel (10 nM). |
Sample Preparation:
| Sampleprep ID: | SP000470 |
| Sampleprep Summary: | Added 1 mL of an ice-cold chloroform to the tubes and 10 ceramic beads. Tubes were vortexed for 30 seconds on a multi-tube vortexer at 5000 rpm three times to homogenize and centrifuged samples at 4 °C in a swinging bucket centrifuge for 60 min at 3,700 rpm. Majority of aqueous (top) layer was carefully transferred to 5 mL cryotubes. Majority of lipid (bottom) layer was carefully transferred to 7 mL glass vials. Remaining protein layer and residual aqueous & lipid layers were transferred into a 2 mL Lo-Bind tubes. Original tubes were rinsed with 600 µL of 2:1 chloroform:methanol solution and transferred to the 2 mL tubes. Centrifuged tubes at 15,000 rpm at 4 °C for 20 min and transferred remaining aqueous & lipid fractions into respective tubes. For each study sample, an 800 µL aliquot of the aqueous fractions was transferred to labeled 2.0 mL Lo-Bind tubes. Analytical pooled QC samples were generated by transferring an additional 75 µL aliquot from all study samples into a 10 mL tube. The total pooled sample was vortexed and 800 µL aliquots were transferred to 2.0 mL tubes labeled Pool. All tubes were frozen for 1 hr and lyophilized to dryness overnight. Samples were reconstituted by adding 700 µL of 90:10 D2O:Chenomx ISTD master mix to each, vortexed for 2 mins and centrifuged at 16,000 rcf for 4 min. A 600 µL aliquot of the supernatant was transferred into 5 mm NMR tubes (Bruker-Biospin, Switzerland), and kept on ice until data acquisition. |
| Processing Method: | lyophilization |
| Extraction Method: | Acetonitrile:water and chloroform |
| Sample Resuspension: | NMR master mix |
| Cell Type: | breast cancer |
Analysis:
| MB Sample ID: | SA022544 |
| Analysis ID: | AN000693 |
| Laboratory Name: | David H. Murdock Research Institute. |
| Analysis Type: | NMR |
| Acquisition Date: | 7/29/14 and 8/13/14 |
| Software Version: | TopSpin 3.2 |
| Operator Name: | Jason Winnike |
| Detector Type: | NMR |
| Num Factors: | 2 |
NMR:
| NMR ID: | NM000077 |
| Analysis ID: | AN000693 |
| Instrument Name: | Bruker Avance III 600 MHz |
| Instrument Type: | FT-NMR |
| NMR Experiment Type: | 1D 1H |
| Field Frequency Lock: | Deuterium |
| Standard Concentration: | 0.5 mM DSS |
| Spectrometer Frequency: | 600 MHz |
| NMR Probe: | 5 mm ATMA Cryoprobe |
| NMR Solvent: | D2O |
| NMR Tube Size: | 5 mm |
| Shimming Method: | Topshim |
| Pulse Sequence: | noesyprid |
| Water Suppression: | yes |
| Pulse Width: | 9.28 us |
| Power Level: | 39.811 W |
| Receiver Gain: | 50.8 |
| Offset Frequency: | 2817.50 Hz |
| Presaturation Power Level: | 0.000025119 W |
| Chemical Shift Ref Cpd: | DSS |
| Temperature: | 298.1 K |
| Number Of Scans: | 256 |
| Dummy Scans: | 4 |
| Acquisition Time: | 2.482 s |
| Relaxation Delay: | 2 s |
| Spectral Width: | 11.0011 ppm, 6602.113 Hz |
| Num Data Points Acquired: | 32768 |
| Real Data Points: | 16384 |
| Line Broadening: | 0.5 Hz |
| Zero Filling: | yes |
| Apodization: | Lorentzian |
| Baseline Correction Method: | Polynomial |
| Chemical Shift Ref Std: | DSS |
| Binned Increment: | 0.04 |
| Binned Data Excluded Range: | water (4.66 - 4.86 ppm); imidazole (7.15-7.26 ppm) |
