Bio-Rad Bio-Gel P Polyacrylamide Gel - Manual (Page 11)

Table 4. Formulations for buffers and solutions provided in the GST
kits.
Profinia Solution Supplied Concentrated Formulation Working 1x Formulation Volume Position #
As
GST lysis buffer 2x 300 mM NaCl, 20 mM Na
2
HP0
4
, 150 mM NaCl, 10 mM Na
2
HP0
4
, 125 ml N/A
10 mM EDTA, pH 7.4 5 mM EDTA, pH 7.4
GST wash buffer 2x 300 mM NaCl, 20 mM Na
2
HP0
4
, 150 mM NaCl, 10 mM Na
2
HP0
4
, 200 ml 1
10 mM EDTA, pH 7.4 5 mM EDTA, pH 7.4
GST elution buffer 2x 40 mM glutathione, 200 mM Tris, 20 mM glutathione, 100 mM 100 ml 3
10 mM EDTA, pH 8.0 Tris, 5 mM EDTA, pH 8.0
Desalting buffer 5x 685 mM NaCl, 13.5 mM KCl, 137 mM NaCl, 2.7 mM KCl, 200 ml 4
21.5 mM Na
2
HP0
4
, 40.5 mM 4.3 mM Na
2
HP0
4
,
KH
2
HP0
4
, pH 7.0 8.1 mM KH
2
P0
4
, pH 7.4
(pH 7.4 upon dilution)
Cleaning solution 1 2x 1,000 mM NaCl, 100 mM Tris, 500 mM NaCl, 50 mM Tris, 125 ml 5
pH 8.0 pH 8.0
Cleaning solution 2 4x 2,000 mM NaCl, 400 mM NaOAc, 500 mM NaCl, 100 mM NaOAc, 125 ml 6
pH 4.5 pH 4.5
Storage solution 2x 4% C
6
H
5
CH
2
OH 2% C
6
H
5
CH
2
OH 200 ml 7
(benzyl alcohol) (benzyl alcohol)
4.2 Instructions for Use
The buffers are provided as concentrates and are diluted to a
1x formulation by the instrument using the preprogrammed methods. The
entire contents (1.23 g) of the glutathione pack need to be added to the
GST elution buffer prior to a purification. Add 5 ml of the elution buffer to
the vial of glutathione and mix to completely resuspend. Either pipet or
pour the glutathione solution into the elution buffer bottle. Filter the solution
through a 0.2 µm filter to remove particulates. The position labeling of the
bottles and insertion into the instrument is identical to that described in
Section 2.2. Note: GST methods do not use a second wash buffer
and position 2 is left empty or filled with water
. Refer to Figure 4 for
proper placement of the GST buffer bottles. If less than 10 ml of purifications
are performed, the glutathione-containing elution buffer can be frozen in
aliquots and used within 3 months. The stored elution buffer should only
be thawed once. The other GST buffers can be capped and stored at 4°C
for up to 3 months.
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