Bio-Rad Experion Protein Analysis Kits - Manual (Page 11)

1.2.4 Prepare the BGG Dilutions
Prepare a serial dilution of the bovine g-globulin (BGG, 2.0 mg/ml) supplied with the kit.
Label three 0.65 ml microcentrifuge tubes 1–3, and add 200 µl DEPC-treated water into
each tube. Then, add the following:
Tube 1 (1,000 ng/µl): 200 µl BGG stock solution
Tube 2 (500 ng/µl): 200 µl BGG dilution from tube 1
Tube 3 (250 ng/µl): 200 µl BGG dilution from tube 2
Notes: You may also use samples 1 and 2 in Test 2 3 . If running samples over multiple
days, store these samples at 4°C between uses.
1.2.5 Prepare the Sample Buffers
Protein samples can be separated under either reducing or nonreducing conditions, but
the Pro260 ladder must always be run under reducing conditions. Reduced and nonreduced
samples may be run on the same chip. Prepare fresh sample buffer each day that an
assay is run.
The starter kit includes dithiothreitol (DTT) in place of the b-mercaptoethanol specified in
the Experion Pro260 analysis kit instruction manual. For the purpose of Experion Pro260
analysis, these reducing agents are interchangeable.
1. Reconstitute the DTT by adding 1 ml DEPC-treated water. This solution remains stable
for weeks after preparation if stored at –20°C.
2. Transfer 30 µl sample buffer (yellow cap) to a 0.65 ml microcentrifuge tube. Add 1 µl of
the reconstituted DTT solution, and label this tube “R” for “reducing”.
3. Transfer another 30 µl sample buffer (yellow cap) to another 0.65 ml microcentrifuge
tube. Add 1 µl DEPC-treated water, and label this tube “NR” for “nonreducing”.
Note: When you are finished preparing the sample buffers, return the stock sample buffer
(yellow cap) to storage at 4°C and the reconstituted DTT to storage at –20°C. Protect all
tubes containing sample buffer from direct sunlight.
3 Do not use these samples for Test 3. For Test 3, prepare the entire dilution series as directed and at the same time for
best results.
7
10010510A.qxp 2/13/2008 11:43 AM Page 7