Summary of Study ST000539
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR000315. The data can be accessed directly via it's Project DOI: 10.21228/M8002N This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST000539 |
| Study Title | Metabolomics-based elucidation of active metabolic pathways in erythrocytes and HSC-derived reticulocytes (part II) |
| Study Type | Cell type comparison |
| Study Summary | Human stem cell derived reticulocytes were compared with mature erythrocytes by metabolomics analysis. |
| Institute | Monash University |
| Department | Monash Institute of Pharmaceutical Sciences, Drug Delivery, Disposition and Dynamics |
| Laboratory | Creek lab |
| Last Name | Creek |
| First Name | Darren |
| Address | 381 Royal Parade, Parkville, Melbourne, VIC3052, Australia |
| Darren.Creek@monash.edu | |
| Phone | N/A |
| Submit Date | 2017-01-23 |
| Num Groups | 6 |
| Total Subjects | 18 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2017-07-10 |
| Release Version | 1 |
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Sample Preparation:
| Sampleprep ID: | SP000568 |
| Sampleprep Summary: | Metabolism was quenched by immersion of cultures in an ethanol/dry ice bath to 0-4 °C. Cells were pelleted by centrifugation (10,000 rpm for 1 min) and washed in cold PBS (1 mL). Metabolites were extracted from 1 x 108 RBCs and 0.5 x 108 reticulocytes by addition of 300 µL chloroform/methanol/water (1:3:1 v/v) containing internal standards (CHAPS, CAPS, PIPES and TRIS; 1 µM) and left for 30 mins at 4 °C with periodic mixing and sonication. The number of cells used for RBCs and reticulocytes was based on the mean cell volume of each cell type, and ensured that the total cell pellet volume was equivalent for each sample, allowing direct comparison of metabolite concentrations from these samples. After mixing, cellular debris was removed by centrifugation (16,000 rpm for 10 mins) and the supernatant was kept at -80 °C prior to analysis. |
| Processing Method: | Lysis with mixing and sonication at 4°C |
| Processing Storage Conditions: | on ice or 4°C |
| Extraction Method: | chloroform/methanol/water (1:3:1 v/v) |
| Extract Storage: | -80°C |
| Sample Spiking: | internal standards (CHAPS, CAPS, PIPES and TRIS; all at 1 µM) mixed in extraction solvent |
| Cell Type: | Human stem cell derived reticulocytes and mature erythrocytes |
