Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA159039

Local Sample ID:	Ctl_61120
Subject ID:	SU001785
Subject Type:	Cultured cells
Subject Species:	Mus musculus
Taxonomy ID:	10090

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Sample Preparation:

Sampleprep ID:	SP001791
Sampleprep Summary:	Protocol for Lipid Extraction (10.24.2019) Lipid Biomarker for Senolysis Study 1. 80% MeOH was added to each sample 500µL to plasma samples 100µL to urine sample 2. Transfer the contents to 9 mL glass tube and add 1.0 mL of CHCl3 (containing 200 ng/mL FA 17:0 Saturated). Vortex mix for 5 minutes. 3. Place the glass tube into a 15mL polypropylene culture tube with Kimi wipes inserted at bottom as a cushion. Wrap the culture tube with paraffin and place in centrifuge. This will prevent glass tube being shattered in the centrifuge. 4. Centrifuge at 4°C for 10 minutes 3000rpm. 5. After spinning there will be two distinct layers, the top layer is the aqueous layer containing the aqueous metabolites (80% MeOH) and bottom layer which contains lipids. 6. Transfer the lower layer into 1.5ml vials. 7. Transfer from the 1.5mL tube the exact amount of each sample into the MS/HPLC glass vial and dry down using nitrogen gas. 8. Reconstitute the lipids with 80µL chloroform. 9. Transfer upper layer (metabolites) into labeled vials and store at -80°C, if possible. Samples stored in -80°C. Sample volume submitted -Plasma samples=50µL Urine samples= 10µL Sample volume in each vial- Plasma samples µL and Urine samples µL -submitted to mass spec All samples were split in half ½ given to CW for further testing other 1/2 in -80°C

Sampleprep ID:

SP001791

Sampleprep Summary:

Protocol for Lipid Extraction (10.24.2019) Lipid Biomarker for Senolysis Study 1. 80% MeOH was added to each sample 500µL to plasma samples 100µL to urine sample 2. Transfer the contents to 9 mL glass tube and add 1.0 mL of CHCl3 (containing 200 ng/mL FA 17:0 Saturated). Vortex mix for 5 minutes. 3. Place the glass tube into a 15mL polypropylene culture tube with Kimi wipes inserted at bottom as a cushion. Wrap the culture tube with paraffin and place in centrifuge. This will prevent glass tube being shattered in the centrifuge. 4. Centrifuge at 4°C for 10 minutes 3000rpm. 5. After spinning there will be two distinct layers, the top layer is the aqueous layer containing the aqueous metabolites (80% MeOH) and bottom layer which contains lipids. 6. Transfer the lower layer into 1.5ml vials. 7. Transfer from the 1.5mL tube the exact amount of each sample into the MS/HPLC glass vial and dry down using nitrogen gas. 8. Reconstitute the lipids with 80µL chloroform. 9. Transfer upper layer (metabolites) into labeled vials and store at -80°C, if possible. Samples stored in -80°C. Sample volume submitted -Plasma samples=50µL Urine samples= 10µL Sample volume in each vial- Plasma samples µL and Urine samples µL -submitted to mass spec All samples were split in half ½ given to CW for further testing other 1/2 in -80°C