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MB Sample ID: SA217782
| Local Sample ID: | 3hpa_G |
| Subject ID: | SU002357 |
| Subject Type: | Other organism |
| Subject Species: | Xenopus tropicalis |
| Age Or Age Range: | Stage 41-43 |
| Species Group: | Amphibians |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002357 |
| Subject Type: | Other organism |
| Subject Species: | Xenopus tropicalis |
| Age Or Age Range: | Stage 41-43 |
| Species Group: | Amphibians |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| 3hpa_G | SA217782 | FL026435 | 3hpa | Timepoint |
Collection:
| Collection ID: | CO002350 |
| Collection Summary: | Tadpoles were anesthetized with 0.05% MS-222 in 1/9x MR and tested for response to touch prior to tissue collection. For uninjured tips (Tips), a sterilized scalpel was used to amputate 500µm of the posterior tip. For 0 hours post amputation (hpa) samples, the posterior third of the tail was amputated followed by collection of tail tissue 250µm anterior to the initial wound. For 3 and 24hpa, 250µm of tissue, including the regenerating tissue, was collected. 10 replicates of 25 tails (8 replicates for the 24hpa timepoint) were collected. For each sample, media was removed, and samples were frozen on dry ice within 5-8 minutes from the first amputation. |
| Sample Type: | Tails |
Treatment:
| Treatment ID: | TR002369 |
| Treatment Summary: | No applicable |
Sample Preparation:
| Sampleprep ID: | SP002363 |
| Sampleprep Summary: | Aqueous metabolites for targeted LC-MS analysis were extracted using a protein precipitation method similar to the one described elsewhere (Mathon et al., 2019; Meador et al., 2020). Samples were first homogenized in 200 µL purified deionized water at 4 ˚C, and then 800 µL of methanol containing 6C13-glucose and 2C13-glutamate (reference internal standards) was added. Afterwards samples were vortexed, stored for 30 minutes at -20 ˚C, sonicated in an ice bath for 10 minutes, centrifuged for 15 min at 14,000 rpm and 4 ˚C, and then 600 µL of supernatant was collected from each sample. Lastly, recovered supernatants were dried on a SpeedVac and reconstituted in 1.0 mL of LC-matching solvent containing 2C13-tyrosine and 3C13-lactate (reference internal standards). |
| Sampleprep Protocol Filename: | JPatel_LC-MS_Methods.pdf |
Combined analysis:
| Analysis ID | AN003712 | AN003713 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | HILIC | HILIC |
| Chromatography system | Shimadzu Nexera X2 | Shimadzu Nexera X2 |
| Column | Waters XBridge BEH Amide XP | Waters XBridge BEH Amide XP |
| MS Type | ESI | ESI |
| MS instrument type | Triple quadrupole | Triple quadrupole |
| MS instrument name | ABI Sciex 6500+ QTrap | ABI Sciex 6500+ QTrap |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | Peak Intensity | Peak Intensity |
Chromatography:
| Chromatography ID: | CH002750 |
| Instrument Name: | Shimadzu Nexera X2 |
| Column Name: | Waters XBridge BEH Amide XP |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS003461 |
| Analysis ID: | AN003712 |
| Instrument Name: | ABI Sciex 6500+ QTrap |
| Instrument Type: | Triple quadrupole |
| MS Type: | ESI |
| MS Comments: | MS data acquisition was performed in multiple-reaction-monitoring (MRM) mode. LC-MS system was controlled using AB Sciex Analyst 1.6.3 software. Measured MS peaks were integrated using AB Sciex MultiQuant 3.0.3 software. The LC-MS assay was targeting 361 metabolites (plus 4 spiked reference internal standards). |
| Ion Mode: | POSITIVE |
| Analysis Protocol File: | JPatel_LC-MS_Methods.pdf |
| MS ID: | MS003462 |
| Analysis ID: | AN003713 |
| Instrument Name: | ABI Sciex 6500+ QTrap |
| Instrument Type: | Triple quadrupole |
| MS Type: | ESI |
| MS Comments: | MS data acquisition was performed in multiple-reaction-monitoring (MRM) mode. LC-MS system was controlled using AB Sciex Analyst 1.6.3 software. Measured MS peaks were integrated using AB Sciex MultiQuant 3.0.3 software. The LC-MS assay was targeting 361 metabolites (plus 4 spiked reference internal standards). |
| Ion Mode: | NEGATIVE |
| Analysis Protocol File: | JPatel_LC-MS_Methods.pdf |
