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Unformatted text preview: 1. We use the protein standard BSA Why do we use BSA? Cheat as dirt Average amino acid composition standard protein 2. Method: making a calibration curve (see ppt slide 2) Known concentration of BSA to brafford reagent.s 3. after making the std curve, you need to assay YOUR fractions and determine [protein] Look at ppt slide 1 Fraction 7 = 0.15 mg/ml Total protein = [protein] mg/ml X vol of fraction (ml) = total protein in mg = 0.15 mg/ml X 1.5 ml = 0.225 mg 4. Enzyme activity (our second assay) Purpose: determine alk phos activity in our fractions (the same ones we used for our Bradford assay) How much activity is there in this fraction tube? See ppt slide 2. The nuts and bolts of the activity assay: Determining total activity A410 /3 (min)/vol assayed (ml) X total vol of fraction (mls) = A410/min (convert A410 to concentration using Beers law)...
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