Return to study ST000361 main page
MB Sample ID: SA016032
| Local Sample ID: | B_0182 |
| Subject ID: | SU000382 |
| Subject Type: | human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Age Or Age Range: | 27-61 years |
| Gender: | male/female |
| Human Race: | caucasian, hispanic, african american |
| Species Group: | Human |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU000382 |
| Subject Type: | human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Age Or Age Range: | 27-61 years |
| Gender: | male/female |
| Human Race: | caucasian, hispanic, african american |
| Species Group: | Human |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| B_0182 | SA016032 | FL003494 | Control | Group |
Collection:
| Collection ID: | CO000376 |
| Collection Summary: | For extraction, 500 µL of an ice-cold solution of 0.005 mg/mL Tryptohan-d5 in 90:10 Methanol:Chloroform (v/v) was added to the tubes containing the thawed human dermal fibroblasts cell pellets. MagNA Lyser ceramic beads were added to the tubes, and a MagNA Lyser was used to beat the samples for two thirty seconds pulses at 2,000 rpm, placing the samples on cold block for 5 minutes in between pulses. Sonication for five minutes was performed to the beaten cell samples, followed by centrifugation at room temperature and at 16,000 rcf for 4 minutes. A 300 µL aliquot of each experimental cell sample homogenized supernatant was transferred to new pre-labeled 2.0 mL LoBind Eppendorf tubes and stored at -80 °C for one hour. |
| Sample Type: | fibroblasts |
Treatment:
| Treatment ID: | TR000396 |
| Treatment Summary: | NA |
Sample Preparation:
| Sampleprep ID: | SP000389 |
| Sampleprep Summary: | Whole study pooled QC samples were created by combining 175 µL aliquot from each of the study samples into a 20 mL scintillation vial. The QC pooled sample was then vortexed for 30 seconds on an analog vortex mixer. Aliquots, 300 µL, were transferred to five new pre-labeled 2.0 mL LoBind Eppendorf tubes and stored at -80 °C for one hour. Next, the supernatants of the study and whole study pooled QC samples, generated above, were dried overnight using a lyophilizer. The residue was reconstituted in 75 µL of reconstitution buffer ( 95:5 Water:Methanol, v/v) and mixed on a multiple tube vortexer for five minutes at 5,000 rpm. Then, the samples were centrifuged at room temperature at 16,000 rcf for four minutes, and the supernatants were transferred to clean pre-labeled autosampler vials for data acquisition. |
Combined analysis:
| Analysis ID | AN000592 | AN000593 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | Reversed phase | Reversed phase |
| Chromatography system | Waters Acquity | Waters Acquity |
| Column | Waters ACQUITY UPLC HSS T3 | Waters ACQUITY UPLC HSS T3 |
| MS Type | ESI | ESI |
| MS instrument type | QTOF | QTOF |
| MS instrument name | Waters Synapt G2 Si QTOF | Waters Synapt G2 Si QTOF |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | intensity | intensity |
Chromatography:
| Chromatography ID: | CH000424 |
| Chromatography Summary: | UPLC-MS spectra were collected for all samples. UPLC was performed on a Waters Acquity UPLC with an Acquity HSS T3 column (2.1x 100mm x 1.8 µm) at 50 °C using the reversed-phase separation. Water with 0.1% formic acid (mobile phase A) and methanol with 0.1% formic acid (mobile phase B) were used as mobile phases and the metabolites were chromatographically separated using a gradient separation: (see the 3. Sheridan-Mice-Microglia Cells_RP-Metadata and Analytical Metadata.xlsx file for the flow gradient). Mass spectroscopy analysis was performed using a Synapt G2 Si ESI-Q-TOF using a 10 µL injection volume. UPLC-MS data were collected over 70-1000 m/z in both positive and negative modes. |
| Methods Filename: | RTI-RCMRC-RP |
| Instrument Name: | Waters Acquity |
| Column Name: | Waters ACQUITY UPLC HSS T3 |
| Flow Rate: | .4ml/min |
| Injection Temperature: | 8 C |
| Solvent A: | 100% water; 0.1% formic acid |
| Solvent B: | 100% methanol; 0.1% formic acid |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS000528 |
| Analysis ID: | AN000592 |
| Instrument Name: | Waters Synapt G2 Si QTOF |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| Ion Mode: | POSITIVE |
| Analysis Protocol File: | RTI-RCMRC-RP-POS |
| MS ID: | MS000529 |
| Analysis ID: | AN000593 |
| Instrument Name: | Waters Synapt G2 Si QTOF |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| Ion Mode: | NEGATIVE |
| Analysis Protocol File: | RTI-RCMRC-RP-NEG |
