User:Paul Rothenberg/Notebook/Pauls Notebook/2014/06/10
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Purification of Mn-Mb
Going to run the Mn-Mb we made through a PD-10 column with H2O. We will run 25 mL of H2O through to equilibrate the column, and let that run through. Following that, we will add 2.5 mL of protein, and let the band collect at the top. Then, 3.5 mL of H2O will be added, and the band collected. Following collection, 25 mL of H2O will be run through, and that procedure will be repeated until we have collected all the protein.
Andrew is working on this, currently. He will collect the excess porphyrin from the column just for retention purposes. We will run a gel to ensure that it has no protein.
I'm continuing the filtration of Co-PPIX. I collected 687.2 mg so far, and I will continue filtering the solution for a higher yield. (The weight of the empty vial was 13.1737 g, and the vial with the first batch of porphyrin was 13.8609 g.
The mass of the new filter, rubber stopper, and filter paper was 88.3150 g.
For reconsitution of ApoMb with CoPPIX, we made an autoclaved a 100 mM Phosphate buffer. 4.463 g of Sodium Phosphate Monobasic and 8.901 g of Sodium Phosphate Dibasic were dissolved in 1 L of H2O, filtered, and autoclaved.