FastScan™ ELISA Cell Extraction Enhancer Solution (50X) #25243
Product Information
Product Usage Information
1X Cell Extraction Buffer: Prepare by diluting FastScan™ ELISA Cell Extraction Buffer (5X) #69905 and FastScan™ ELISA Cell Extraction Enhancer Solution (50X) #25243* to 1X with deionized water. This buffer can be stored at 4°C for short-term use (1-2 weeks). To make 10 mL 1X Cell Extraction Buffer, combine 7.8 mL deionized water, 2 mL FastScan™ ELISA Cell Extraction Buffer (5X), and 200 μL FastScan™ ELISA Cell Extraction Enhancer Solution (50X). Alternatively, Enhancer Solution may be added to the Cell Extraction Buffer after extraction of cells or tissue. When using the 1X Cell Extraction Buffer as a sample diluent for the assay, it is recommended to equilibrate it to room temperature prior to use.
*IMPORTANT: The provided FastScan™ ELISA Cell Extraction Enhancer Solution (50X) may precipitate when stored at 4°C. To dissolve, warm briefly at 37°C and mix gently. The FastScanTM ELISA Cell Extraction Enhancer Solution (50X) can be stored at room temperature to avoid precipitation.
NOTE: The 1X Cell Extraction Buffer contains phosphatase inhibitors. Protease inhibitors should be added to the 1X Cell Extraction Buffer immediately prior to lysing cells. Additional phosphatase inhibitors can also be added (e.g. Protease/Phosphatase Inhibitor Cocktail (100X) #5872, not supplied).
Preparing Cell Lysates
For adherent cells
1. Aspirate media when the culture reaches 80–90% confluence.
2. Remove media and rinse cells once with ice-cold 1X PBS.
3. Remove PBS and add 0.5 mL ice-cold 1X Cell Extraction Buffer (recommended to supplement with protease inhibitors and additional phosphatase inhibitors as needed) to each plate (10 cm diameter) and incubate the plate on ice for 5 min.
4. Scrape cells off the plate and transfer to an appropriate tube. Keep on ice.
5. Sonicate lysates on ice.
6. Microcentrifuge for 5 min (x14,000 rpm) at 4°C and transfer the supernatant to a new tube. The supernatant is the cell lysate. Store at -80°C in single-use aliquots.
For suspension cells
1. Remove media by low speed centrifugation (~1200 rpm) when the culture reaches 0.5–1.0 x 106 viable cells/ml.
2. Wash once with ice-cold 1X PBS.
3. Cells harvested from 50 mL of growth media can be lysed in 2.0 mL of 1X Cell Extraction Buffer (recommended to supplement with protease inhibitors and additional phosphatase inhibitors as needed).
4. Sonicate lysates on ice.
5. Microcentrifuge for 5 min (x14,000 rpm) at 4°C and transfer the supernatant to a new tube. The supernatant is the cell lysate. Store at -80°C in single-use aliquots.
Storage
Product Description
Limited Uses
Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.
Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.