plate. Turn off vacuum.
16. Add 600 µL of Wash Solution 1 and apply vacuum
until all the solution passes through the binding
plate. Turn off vacuum.
17. Add 600 µL of Wash Solution 2 and apply vacuum
the supernatant.
6. Resuspend the cell pellet in 1 ml of 1× Buffer B.
7. Add 10 µL of the resuspended cell sample to
990 µL of ultrapure water and read the OD at
fluorescent
DNA stains
600 µL vial DPLUS-100HS
DirectLoad™ Plus
1 Kb DNA Ladder
500 µL vial DPLUS-1K
DirectLoad™ Plus
1 Kb DNA Ladder
with fluorescent
DNA stains
600 µL vial DPLUS-1KS
Place spin column back in
this same collection tube.
6. Load the prepared cell extract on the column.
The column will hold up to 600 µL of extract at
one time.
7. Centrifuge
cleared by mixing
600 µL milk with 100 µL 6 N HCl.
2. Centrifuge for 5 minutes at 14,000 × g.
3. Transfer 300 µL supernatant into a clean tube
and neutralize with 50 µL
to remove lipids, which may be present
in the tissue.
6. Transfer the homogenate to a 2 mL Eppendorf
tube. Centrifuge the sample at 600 × g for
5 minutes.
7. Carefully transfer the
unbound
protein from the filter plate using 600 µL of Wash
Buffer with centrifugation. Empty the collection
plate.
9. Repeat the wash step (Step 8) with 600 µL of
Wash Buffer.
10.
in the dark for 15 minutes.
6. Spin down cells at 600 x g for 3 minutes and discard buffer.
7. Resuspend cells in 1 mL of 1X Wash Buffer and spin down cells at 600 x g
tube tightly and place in a 400-600 mL glass beaker filled with approximately 200 mL
of water.
5. Microwave beaker with tube until water in beaker begins boiling.
6. Remove beaker and vortex tube
80500758 968
D51XW300WX4-EU 300 kg 20 g 600 x 600 mm 80251810 1071
D51XW300WX4EU-M 300 kg 100 g 600 x 600 mm 80251820 1129
D51XW300WX4EUMB 300 kg 100 g 600 x 600 mm 80500759 1238
RE-
CHARGE
APPM
mM Tris-HCl (pH 7.5), 10 mM MgCl2, 22 mM
NH4Cl, 10 mM β-mercaptoethanol, and 5% glyercol.6
Elution of protein may be affected by adding NaCl
from 100 mM up to 600 mM to this buffer.3 Removing
tightly