Rat Brain Whole Tissue Lysate (Adult Whole)

Rat

Western Blot

Ready-to-use lysates are especially prepared as positive controls for separation by SDS-PAGE and subsequent western blot analysis. Lysates are prepared in denaturing buffer WITHOUT dissociating agents (i.e. no 2-mercaptoethanol or dithiothreitol has been added). Heat lysate to 95C for 5 minutes and rapidly cool. If dissociating conditions are desired, add reducing agent prior to heating. The recommended loading volume per lane is 10-20 ul depending on the size format of your gel.

Tissue

Adult

Whole

Tissues were washed exhaustively with PBS to remove blood and other debris. A lysate was prepared by homogenizing the tissue and washing the cells in cold PBS. Washed cells were incubated at 4C in modified RIPA buffer containing 150 mM sodium chloride, 50 mM Tris Cl, pH 7.4, 1 mM EDTA, 1.0% NP-40, 0.5% sodium deoxycholic acid and 0.1% SDS to lyse the cells. Protein integrity is ensured using a cocktail of protease inhibitors with broad specificity for the inhibition of aspartic, cysteine, and serine proteases as well as aminopeptidases (0.1 mM AEBSF HCl, 0.08 uM Aprotinin, 5 uM Bestatin, 1.5 uM E-64, 2 uM Leupeptin Hemisulfate and 1 uM Pepstatin A). The following phosphatase inhibitors were also added: 1 mM NaF and 1 mM Na3VO4. Cell debris was removed by centrifugation and membrane filtration. Protein concentration was determined by Lowry assay using a commercially available kit. The protein concentration was adjusted to 2 mg/ml and then an equal volume of 2X SDS-PAGE sample buffer was added.

Multi-step

1X SDS-PAGE Sample Buffer (62.5 mM Tris HCl, 2% SDS, 10% Glycerol and 0.005% bromophenol blue, pH 6.8), 10% (v/v) Glycerol

No Preservative

Please contact technical services for concentration.

The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.

Store at -70C. Avoid freeze-thaw cycles.