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Step 2, First Ammonium Sulfate Fractionation Solid ammonium sulfate was added to the cell-free extract to give 56% saturation (at0℃). The pH was maintained at 6.9 with 2 N KOH. After stirring for 1 h and removal of the precipitate by centrifugation, the supernatant solution was further saturated with ammonium sulfate to give 72% saturation (0°C). After I h, the suspension was centrifuged, and the pellet was dissolved in Buffer A containing 1.25 M ammonium sulfate and 0.03% sodium azide. |

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