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Tibshirani R, Walther G, Botstein D, Brown P (2005)

Supernatants may be isolated by centrifugation at appropriate speed and duration (e.g., 13,000 rpm for 1 min, 500 rpm for 5 min, etc.)

Review your storage conditions: ensure the refrigerator maintains 28C consistently (verify with a thermometer, not the appliance display) and that you're using fresh alcohol swabs for every needle puncture

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And a cell-based bioassay, receptor autophosphorylation or a proliferation readout in a responsive line, is the only method that answers the question you actually care about, which is whether the molecule works

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