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For protein isolation from tissue, the epidermis was isolated as described above.
The brain was removed and the infarcted area isolated and processed for RNA and protein isolation.
Spheroids were isolated from matrix using iso-osmolar trisodium citrate for lysis and protein isolation.
For protein isolation, tissues were processed immediately as described below.
Membrane protein isolation is a challenging problem.
J.H. performed the experiments by assisting with tissue collection; DNA, RNA and protein isolation; and analyses.
Protein isolation was performed by cell lysis using RIPA buffer (Sigma-Aldrich) in the presence of protease inhibitors (Roche).
For high purity sorts required for metabolite profile or protein isolation, neutrophils were further purified by fluorescent associated cell sorting by Ly6G+, purities obtained were >95%.
This block (comprised primarily of the hypothalamus) was immediately wrapped with sterile foil and stored at −80 °C until protein isolation.
Disruption of D. tyermannii leaves using the P-PER™ Plant Protein Extraction Reagent was chosen for routine protein isolation according to the manufacturer's protocol.
To examine reproducibility of the technology, we performed each assay with a technical replicate and tested two different protein isolation platforms (Affymetrix and Signosis).
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