Bio-Rad Experion Protein Analysis Kits - Manual (Page 10)

• Keep the BGG standard on ice
• Equilibrate the following to room temperature for 15–20 min:
–
Dithiothreitiol (DTT)
– Pro260 sample buffer (yellow cap)
– Pro260 ladder (red cap)
– 2 Tubes of Pro260 gel 2 (green cap)
– Pro260 stain (blue cap) 2
Notes: Keep the Pro260 stain (blue cap) and sample buffer (yellow cap) covered (for
example, with foil) at all times to avoid exposure to light.
Inspect the reagents; if the Pro260 gel (green cap) has been frozen, do not use it.
Do not store reagents at room temperature for more than 2 hr, as this will affect their shelf
life.
3. Vortex the contents of each tube and briefly centrifuge to collect the solutions at the
bottoms of the tubes. Make sure that the Pro260 stain solution (blue cap) is completely
thawed before proceeding.
1.2.3 Filter the Gel and Prepare the Gel-Stain Solution
Notes: The Experion Pro260 starter kit includes 2 spin filters, which are sufficient for
preparation of one filtered gel (G) and one gel-stain solution (GS). You will use the G and
GS in Tests 2 and 3. The prepared G and GS can be used for up to 4 weeks when stored
in the dark at 4°C between each use. The solutions can be refiltered once after that to
extend their use for another 4 weeks.
Additional spin filter tubes and reagents may also be purchased separately (see Appendix D
for ordering information).
Prepare GS as described below. Do not prepare smaller aliquots, as doing so may
adversely affect the gel-to-stain ratio, which is critical to successful staining.
1. Prepare the GS by adding 20 µl Pro260 stain (blue cap) to a tube of 520 µl Pro260
gel (green cap). (Each tube of Pro260 gel contains 520 µl gel.) Vortex the GS for
10 sec at the highest setting and then spin it down in a microcentrifuge for a few
seconds.
Note: Cap the Pro260 stain tightly (it contains DMSO, which is highly hygroscopic), and
store the tube in the dark (for example, cover it with foil) at 4°C.
2. Transfer the GS to a spin filter. Label the tube with “GS” and the date.
3. Pipet the contents (520 µl) of the other tube of Pro260 gel (green cap) into another
spin-filter tube. Label the tube with “G” and the date.
4. Centrifuge both spin filters for 5 min at 10,000 × g.
5. Inspect the tubes to ensure that all of the gel has passed through the filters and then
discard the filters.
6. Keep both prepared G and GS at room temperature and covered until ready for use.
2 If the filtered gel (G) and gel-stain solution (GS) were prepared previously, remove these from storage and equilibrate
them to room temperature. Do not remove the Pro260 stain.
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