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Spent medium samples were diluted 1 5 in EIA buffer before being assayed.
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Buffer controls with no added inhibitors were assayed in parallel.
[H]NE transport was assayed in KRH buffer.
Cell extracts were diluted 1 : 20 in assay buffer before being added to the fluorescent substrate as experimentation showed this to be the optimum dilution to minimise the effect of the extraction buffer itself on the fluorescent BODIPY® substrate.
In both cases the resulting fine suspensions were diluted 30-fold in the homogenization buffer before treatments and assays, and they were thoroughly re-suspended by vortex mixing as each aliquot was transferred to the reaction tubes.
The lysate was incubated on ice in immunoprecipitation assay buffer for 2 h before being homogenized using a mortar and pestle.
Samples were then assayed for concentration before being diluted into sample buffer.
Supernatants were then concentrated and exchanged with buffer before assaying on CMC for 1 h or on Avicel for 24 h at 65 and 75 °C.
All syringes were flushed with N2 before being used to transfer buffer, reagents, or proteins from the stock vials to the assay cuvettes.
Tissues were homogenized in phosphatase inhibitor containing solubilization buffer, before protein assay and immunoblot analysis.
Dry extract was resuspended in phosphate buffer immediately before the assays.
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