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MB Sample ID: SA266658
| Local Sample ID: | S_235 |
| Subject ID: | SU002794 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002794 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| S_235 | SA266658 | FL034911 | Study sample | Sample Type |
| S_235 | SA266658 | FL034911 | 3 | Batch |
Collection:
| Collection ID: | CO002787 |
| Collection Summary: | N/A |
| Sample Type: | Blood (whole) |
| Collection Method: | Dried Blood Spot |
| Storage Conditions: | -80℃ |
Treatment:
| Treatment ID: | TR002803 |
| Treatment Summary: | NA |
Sample Preparation:
| Sampleprep ID: | SP002800 |
| Sampleprep Summary: | 279 newborn dried blood spot and blank samples and were randomized and extracted by 1000 µL methanol with 500 ng/mLtryptophan-d5 as internal standard via a multiple tube vortex mixer for 10 minutes at 5000 rpm. The blank and study samples were sonicated in an ice bath for 20 minutes. A volume of 700 µL of the supernatants was transferred into a pre-labeled 2.0 mL Low-bind Eppendorf tube labeled as the stock. Protein precipitate was pelleted using a centrifuge operating at 4° C for 5 minutes at 16,000 rcf. A volume of 70 µL of each blank sample stock was transferred to 15 mL conical tube to be combined to create a Blank Pool (BP). A volume of 70 µL of each study sample stock was transferred to 15 mL conical tube to be combined and create a Study Pool (SP). A total volume of 1400 µL of NIST Reference Plasma (NIST) was aliquoted into twenty-eight 50 µL aliquots. A volume of 600 µL supernatant (including experimental samples, BP, and SP) was transferred into a pre-labeled 2.0 mL Low-bind Eppendorf tube. All samples and pools were then dried by a SpeedVac overnight. For immediate analysis, 100 µL of water-methanol solution (95:5, v/v) was added to reconstitute the dried extracts, and the samples were thoroughly mixed on multiple tube vortex mixer for 10 min at 5000 rpm. Samples were centrifuged at 4°C for 4 min at 16,000 rcf. The supernatant was transferred to pre-labeled autosampler vials for data acquisition by LC-MS. Broad-spectrum metabolomics was conducted using a Thermo Scientific™ Vanquish™ UPHPLC - Q Exactive™ HF-X Orbitrap System. Metabolites were separated on an Acquity UPLC HSS T3 C18 (2.1 X 100 mm, 1.8 µm) operating at 50 C using a reversed phase gradient separation with Water with 0.1% Formic Acid (v/v) as mobile phase A and Methanol with 0.1% Formic Acid (v/v) as mobile phase B. A 5 µL was injected into the instrument, and MS was collected between 50-750 m/z in positive mode with the MS/MS data triggered by the Data-Dependent Acquisition. |
| Processing Storage Conditions: | 4℃ |
| Extraction Method: | Vortex with methanol containing 500ng/ml tryptophan-d5 as internal standard |
| Extract Storage: | -80℃ |
| Sample Resuspension: | Water-Methanol (95:5, v/v) |
| Sample Spiking: | Tryptophan-d5 stock solution at 500 ng/mL |
Combined analysis:
| Analysis ID | AN004365 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Thermo Vanquish |
| Column | Waters ACQUITY UPLC HSS T3 (100 x 2.1mm,1.8um) |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive HF-X Orbitrap |
| Ion Mode | POSITIVE |
| Units | Normalized intensity |
Chromatography:
| Chromatography ID: | CH003271 |
| Chromatography Summary: | Reverse phase |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Waters ACQUITY UPLC HSS T3 (100 x 2.1mm,1.8um) |
| Column Pressure: | 6000-10000 psi |
| Column Temperature: | 50℃ |
| Flow Gradient: | Time(min) Flow Rate %A %B Curve 1. 0 0.4 99.0 1.0 5 2. 1.00 0.4 99.0 1.0 5 3. 16.00 0.4 1.0 99.0 5 4. 19.00 0.4 1.0 99.0 5 5. 19.50 0.4 99.0 1.0 5 6. 22.00 0.4 99.0 1.0 5 |
| Flow Rate: | 0.4 mL/min |
| Injection Temperature: | 8℃ |
| Internal Standard: | Tryptophan-d5 |
| Solvent A: | 100% water; 0.1% formic acid |
| Solvent B: | 100% methanol; 0.1% formic acid |
| Analytical Time: | 22 min |
| Weak Wash Solvent Name: | 10% methanol/90% water; 0.1% formic acid |
| Strong Wash Solvent Name: | 75% 2-Propanol/25% Water; 0.1% formic acid |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS004112 |
| Analysis ID: | AN004365 |
| Instrument Name: | Thermo Q Exactive HF-X Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Instrument: Thermo Q Exactive HFx Software: Xcalibur 4.1.31.9 for data acquisition; Progenesis QI 2.4 for data preprocessing |
| Ion Mode: | POSITIVE |
| Capillary Temperature: | 320 °C |
| Capillary Voltage: | 3.5 KV |
| Collision Energy: | 20-45, ramp |
| Collision Gas: | N2 |
| Dry Gas Flow: | 55 |
| Dry Gas Temp: | 400°C |
| Fragmentation Method: | CID |
| Ion Spray Voltage: | 3.5kV |
| Ionization: | ES+ |
| Mass Accuracy: | 5 ppm |
| Dataformat: | Profile |
| Desolvation Gas Flow: | 55 |
| Desolvation Temperature: | 400 ℃ |
