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MB Sample ID: SA252370
| Local Sample ID: | 15mins-1 |
| Subject ID: | SU002607 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002607 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| 15mins-1 | SA252370 | FL032080 | Wild-type | Genotype |
| 15mins-1 | SA252370 | FL032080 | 15mins | Treatment |
Collection:
| Collection ID: | CO002600 |
| Collection Summary: | The eye organoids were collected and washed twice with PBS before pellets were flash-frozen and stored at −80°C until metabolite extraction. Three biologically independent repeated samples were collected. |
| Collection Protocol Filename: | Glucose_and_lactate_isotope_labeling.pdf |
| Sample Type: | Retina |
Treatment:
| Treatment ID: | TR002619 |
| Treatment Summary: | All the organoids (50 organoids per condition) were washed with blank SILAC before being reconstituted in 13C medium at a concentration of 10 eye organoids/ml The incubation was performed from 15 minutes to 2 hours for time-course analysis of the glucose labeling. |
Sample Preparation:
| Sampleprep ID: | SP002613 |
| Sampleprep Summary: | The eye organoids were collected and washed twice with PBS before pellets were flash-frozen and stored at −80°C until metabolite extraction. Metabolite analysis was performed as described previously (PMID: 33931446). Briefly, for metabolite extraction, 80% methanol was used followed by the rapid freeze-thaw method to break the tissues. The supernatant underwent speedvac drying. The samples were prepared in 80% acetonitrile and were analyzed by High-Resolution Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS). |
Combined analysis:
| Analysis ID | AN004129 |
|---|---|
| Analysis type | MS |
| Chromatography type | HILIC |
| Chromatography system | Q-exactive |
| Column | Waters XBridge BEH Amide (100 x 3.0mm, 3.5um) |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive Orbitrap |
| Ion Mode | UNSPECIFIED |
| Units | Peak area |
Chromatography:
| Chromatography ID: | CH003059 |
| Instrument Name: | Q-exactive |
| Column Name: | Waters XBridge BEH Amide (100 x 3.0mm, 3.5um) |
| Column Temperature: | 40 |
| Flow Gradient: | 0 min, 15% A; 2.5 min, 30% A; 7 min, 43% A; 16 min, 62% A; 16.1-18 min, 75% A; 18-25 min, 15% A with a flow rate of 150 μL/min |
| Flow Rate: | 150 μL/mi |
| Solvent A: | 95% water/5% acetonitrile; 10 mM ammonium hydroxide; 10 mM ammonium acetate, pH 9.0 |
| Solvent B: | 100% acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS003876 |
| Analysis ID: | AN004129 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The capillary of the ESI source was set to 275 °C, with sheath gas at 35 arbitrary units, auxiliary gas at 5 arbitrary units and the spray voltage at 4.0 kV. In positive/negative polarity switching mode, an m/z scan range from 60 to 900 was chosen and MS1 data was collected at a resolution of 70,000. The automatic gain control (AGC) target was set at 1 × 106 and the maximum injection time was 200 ms. The top 5 precursor ions were subsequently fragmented, in a data-dependent manner, using the higher energy collisional dissociation (HCD) cell set to 30% normalized collision energy in MS2 at a resolution power of 17,500. Besides matching m/z, metabolites are identified by matching either retention time with analytical standards and/or MS2 fragmentation pattern. Data acquisition and analysis were carried out by Xcalibur 4.1 software and Tracefinder 4.1 software, respectively (both from Thermo Fisher Scientific) |
| Ion Mode: | UNSPECIFIED |
