Bio-Rad Rotofor® and Mini Rotofor Cells - Manual (Page 12)

5. Mount the needle array on the two alignment pins on the bottom of the
chamber. Grasp the needle array with the fingers of both hands while plac-
ing the thumbs on the top of the focusing chamber. Take care not to block
any of the uppermost ports. Quickly push the needles firmly and uniformly
all the way through the sealing tape into the chamber. This will cause all 20
fractions to be simultaneously aspirated from the cell and delivered to the
collection tubes.
Fig. 7. Harvesting samples after focusing is complete. Make sure thumbs do not
cover the uppermost ports.
6. Turn off the vacuum source and remove the test tube rack. Note that all the
odd numbered fractions are in one row and the even numbered fractions
are in the other row of the rack.
Step 9: Expected Results
1. The sample separated was a mixture of three naturally colored proteins in
distilled water containing 2.0% pH 3-10 Bio-Lyte ampholytes.
A. Phycocyanin consists of three blue protein subunits of pI 4.5, 4.7, and 5.0.
These should focus nearest the anode in 1-2 fractions.
B. Hemoglobin A and Hemoglobin C are two red colored proteins of pI
7.1 and 7.4, respectively. These should focus near the middle of the
focusing chamber in 2-3 fractions.
C. Cytochrome c is an orange protein with a pI of 9.6. This protein should
focus near the cathode in 1 to 2 fractions.
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LIT603B 7/10/98 11:58 AM Page 10