ELISA Lysis and Protein Extraction Buffer

N° du produit ABIN1059067

Détail du produit

Application: Protein Extraction (PEx), ELISA

Attributs du produit: Validated formulation for ELISA
Convenient pre-mixed 1X solution
Compatible with protein concentration assays such as Bradford
Widely applicable for extraction of most proteins and glycoproteins from cells.
Also provided: 100X broad spectrum phosphatase inhibitor in a separate tube

Ingrédients:

• 60 mL ELISA protein extraction buffer solution
• 1 mL 100 X Broad Spectrum Phosphatase Inibitor (store at -20 °C)

Information d'application

Commentaires:

This ELISA protein extraction buffer contains a mild detergent enabling desolution of cell membranes, solubilization of most membrane proteins and suspension of cytosolic proteins into the buffer. A separate vial containing a 100X broad spectrum phosphatse inhibitor cocktail is provided to preserve seryl/threonyl and tyrosyl phosphorlation when needed.

Procédure de l'essai:

1. Prepare 500 µl ELISA protein extraction buffer for each 10-cm dish, or 200 µl for each well in a 6-well dish. Make the solution ice cold and add protease inhibitors, including serine protease inhibitor (such as PMSF). If needed, also aliquot provided phosphatase inhibitor which is provided 100X.
   2. Remove media and rinse cells three times with PBS.
   3. Add 500 µl ELISA protein extraction buffer into each 10-cm dish. Rotate the dish to cover cells with ELISA protein extraction buffer and place on a bed of ice for 5 min.
   4. Scrape cells off the dish, and transfer to an appropriate tube. Rotate and tap the tube several times to solubilize membranes. You should observe fibrous materials which correspond to genomic DNA, an indicator of cell lysis. If fibrous materials are not apparent, add 20 µl aliquots of ELISA protein extraction buffer, and rotate and tap until the fibrous materials are observed. Keep the tube on ice for another 15 min.
   5. Centrifuge the tube at 4oC for 15min and collect the supernatant, which will be used in the ELISA assay. A protein concentration assay, such as the Bradford Assay, can be used to quantify total protein concentration. The supernatant can also be stored at -80oC long term.

Restrictions: For Research Use only

Stockage

Format: Liquid

Stock: 4 °C

Référence

Huang, Cheng, Su, Lin: "Syndecan-4 Promotes Epithelial Tumor Cells Spreading and Regulates the Turnover of PKC? Activity under Mechanical Stimulation on the Elastomeric Substrates." dans: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology, Vol. 36, Issue 4, pp. 1291-1304, (2015) (PubMed).