Frequently asked questions
Suggested extraction procedures For research use only. Not for use in diagnostic procedures.
How do I handle Avanti Mass Spec Standards properly? Your Avanti Mass Spec Standard should be stored in a -10°C to -25°C freezer until ready for use. It is designed to be a one-time use ampule, and we do not recommend extended storage after opening. Always make sure to warm bath sonicate the unopened ampoule for approximately 2 minutes prior to opening the ampoule. Lipids in solution may precipitate during shipping and storage conditions, and it may not be visible with a solution at extremely low concentrations such as this. Directly transfer from ampule to sample preparation glass vial for immediate use. General handling guidelines for lipids should be followed as outlined on our website. How do I order Avanti Mass Spec Standards? Please visit the Avanti website. Customers in the United States can order directly from Avanti, and customers outside the United States will be directed to our worldwide distribution partner for country specific ordering information and pricing. Can I use my Avanti Mass Spec Standard more than once? Avanti quantitative mass spectrometry standards are designed to be “one-time use” items. It is important to directly transfer from the ampule to the experiment or prepare as a dilution for immediate use. Avanti cannot guarantee product purity and subsequent performance if used outside these guidelines. How do I reference an Avanti Mass Spec Standard in my publication? Please list the full name and product number of the Avanti product in your materials and methods sections, along with “Avanti Research , Alabaster, AL” as the source. If you publish using an Avanti product we would love to hear about it! Please drop us an email and share your success stories.
How do I open the sealed glass ampule? Ampules are pre-scored for easy opening. With the ampule upright, hold the ampule in a gloved hand and firmly push the top section away from you with even pressure. The top should cleanly snap off, but proceed with care, as it will leave a sharp glass edge. Can I get a Custom Mass Spec Standard? Yes, we custom formulate mass spec standards using the lipids and solvents of your choice. We work with you to design your custom mass spec standard and can make recommendations if needed. Let us know how we can help by emailing lipidomics@avantilipids.com. Avanti has been making Custom Quantitative Mass Spec Standards for the Lipidomics community for over 15 years. We specialize in high-accuracy quantitative packaging of lipids for use as mass spec standards, robotic assisted packaging of solutions, accurate quantitation of lipid concentrations, and rigorous quality control testing. Who do I contact if I have additional questions? Please e-mail us at lipidomics@avantilipids.com if you have any additional questions. Custom Synthesis Avanti prides itself on being innovative by discovering new lipids for our customers. We are constantly adding to our extensive list of products for research. However, if you cannot find the perfect lipid to make your project a success, or you have a suggestion for a new product that is needed in the field, our synthesis team is ready to take on a new challenge. With over 150 years of combined lipid synthesis experience, our team can solve most synthesis problems and deliver custom lipids with Avanti’s signature quality. Please contact our synthesis team at customsynthesis@avantilipids.com to discuss the amazing things we can do for you.
Extraction Protocol for Plasma Using SPLASH TM or UltimateSPLASH™ ONE 1. Use 13 x 100 mm new glass screw capped tubes. Do not use washed tubes as you may extract detergent residue. 2. Add 990 μl water to 10 μl plasma, then let sit on ice for 10 minutes. 3. Add 2.0 mL methanol. 4. Add 0.9 mL dichloromethane. 5. Vortex. 6. A singe phase should appear. If there are two distinct phases, add 50 μl methanol and vortex. If solution is still not a single phase, repeat addition of 50 μl methanol and vortex. 7. Add 10 μl UltimateSPLASH™ ONE Internal Standard, vortex, and let mixture sit for 30 minutes at room temperature. 8. Add 1 mL water. 9. Add 0.9 mL dichloromethane.
4. For cells in suspension: centrifuge, discard saline, and add 1 mL water. Vortex and transfer to glass tube for extraction. Rest on ice for 10 minutes. Ensure final volume is 1 mL and adjust if necessary. 5. For adhered cells: wash cells with non-buffered saline. Add 1 mL water to lyse cells and scrap. Collect cell lysate and transfer to glass tube for extraction. Rest on ice for 10 minutes. Ensure final volume is 1 mL and adjust if necessary. 6. Add 2.0 mL methanol. Add 990 μl water to 10 μl plasma, then let sit on ice for 10 minutes. 7. Add 0.9 mL of dichloromethane. 8. Vortex. 9. Repeat steps 6-16 from Extraction Protocol for Plasma. Extraction Protocol for Cell Culture 1. Weigh tissue to be extracted. 50-100 mg is sufficient. Calculate water content. Expected values are as follows: a.Adipose 18% b. Brain 60% c. Bone 44% d. Heart, kidney, liver, lung, intestines, spleen, and stomach 65% e.Testes 18% 2. Add water to tissue so that total volume is 1 mL. Example: 100 mg of brain tissue corresponds to 60 μL water. Add 940 μL water. 3. Disperse tissue. a. Grind tissue frozen in liquid nitrogen using cold mortar and pestle. b. Blend using a homogenizer. 4. Sonicate for 30 seconds with 5 seconds bursts and 20 second rest time. Perform sonication steps on ice. 5. Add 2.0 mL of methanol. 6. Add 0.9 mL dichloromethane. 7. Vortex. 8. Repeat steps 6-16 from Extraction Protocol for Plasma.
10.Invert tube 10 times. DO NOT VORTEX. 11.Centrifuge at 1200 rpm for 10 minutes.
12.Collect lower layer and put into a new glass tube. 13.Add 2 mL dichloromethane to remains in extraction tube. 14.Mix, centrifuge, and collect lower layer. Add to first extract. 15.Evaporate solvent under a stream of nitrogen. 16.Re-suspend lipids in injection solvent. Extraction Protocol for Cell Culture 1. Use 13 x 100 mm new glass screw capped tubes. Do not use washed tubes as you may extract detergent residue. 2. Collect cells. 3. Wash cells with non-buffered saline to remove cell culture
www.avantiresearch.com
16
17
Made with FlippingBook - professional solution for displaying marketing and sales documents online