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MB Sample ID: SA128430
| Local Sample ID: | Cell_9312_A |
| Subject ID: | SU001598 |
| Subject Type: | Other |
| Subject Species: | Prochlorococcus marinus str. MIT 9312;Prochlorococcus marinus MIT9313 |
| Taxonomy ID: | 74546;74547 |
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Sample Preparation:
| Sampleprep ID: | SP001606 |
| Sampleprep Summary: | Each sample was extracted using a modified Bligh-Dyer extraction. Briefly, quantitative aliquots of cell pellets were transferred into 15 mL teflon centrifuge tubes containing a mixture of 100 µm and 400 µm silica beads. Quantitative aliquots of extracellular vesicles were transferred into 24 mL glass vials and extracted without bead beating. Heavy isotope-labeled internal standards were added along with ~2 mL of cold aqueous solvent (50:50 methanol:water) and ~3 mL of cold organic solvent (dichloromethane). The samples were shaken on a FastPrep-24 Homogenizer for 30 seconds and chilled in a -20 °C freezer repeatedly for three cycles of bead-beating and a total of 30 minutes of chilling. The organic and aqueous layers were separated by spinning samples in a centrifuge at 4,300 rpm for 2 minutes at 4 °C. The aqueous layer was removed to a new glass centrifuge tube. The remaining organic fraction was rinsed three more times with additions of 1 to 2 mL of 50:50 methanol:water. All aqueous rinses were combined for each sample and ~2 mL of cold dichloromethane was added to the combined aqueous layer. Tubes were shaken and centrifuged at 4,300 rpm for 2 minutes at 4°C. The aqueous layer was removed to a new glass vial and dried under N2 gas. The remaining organic layer in the bead beating tubes was transferred into the glass centrifuge tube and the bead beating tube was rinsed two more times with cold organic solvent. The combined organic rinses were centrifuged, transferred to a new glass vial, and dried under N2 gas. Dried aqueous fractions were re-dissolved in 380 µL of water. Dried organic fractions were re-dissolved in 380 µL of 1:1 water:acetonitrile. 20 µL of isotope-labeled injection standards in water were added to both fractions. Process blanks (MilliQ water), media blanks, and PBS (vesicle suspension buffer) were extracted and analyzed alongside each sample set. |
| Processing Storage Conditions: | On ice |
| Extraction Method: | Bligh-Dyer |
| Extract Storage: | -80℃ |
