Clontech Laboratories His60 Ni Superflow Cartridges Benutzerhandbuch - Seite 7
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III.
Additional Materials Required
A. Equipment
For His60 Ni Superflow Cartridges, you will need the following equipment:
A suitable liquid chromatography system (LC procedure only) such as ÄKTA or other systems
A syringe or manual pump
NOTE: For best results, process all buffers through a 0.45 μm filter and degas before use in LC applications.
B. Buffers – Native Conditions
For your convenience, we provide a separate kit containing a set of His60 Ni Extraction, Equilibration, and
Elution Buffers, the His60 Ni Buffer Set (Cat. No.635665), which is sufficient for approximately 20 purifications
on 1 ml His60 Ni Superflow Cartridges (5 x 1 ml) (Cat. No.635675), or 5 purifications on 5 ml His60 Ni
Superflow Cartridges (5 x 5 ml) (Cat. No. 635679).
The His60 Ni Buffer Set is also available together with the 1 ml cartridges in the His60 Ni Superflow Cartridge
Purification Kit (5 x 1 ml) (Cat. No. 635674), and together with the 5 ml cartridges in the His60 Ni Superflow
Cartridge Purification Kit (5 x 5 ml) (Cat. No. 635678)—see Section II.
The following information is provided if you wish to prepare your own buffers for use with other applications.
Please note that for FPLC and other automated applications, you need to filter the buffers through a 0.45 μm filter
and degas them before use.
Equilibration Buffer: 50 mM sodium phosphate, 300 mM sodium chloride, 20 mM imidazole; pH 7.4
Wash Buffer: 50 mM sodium phosphate, 300 mM sodium chloride, 40 mM imidazole; pH 7.4
–– Wash Buffer is easily made on a binary pump LC system by mixing 7.1 parts of His60 Ni Elution
Buffer and 92.9 parts of His60 Ni Equilibration Buffer. This buffer ratio can be achieved by running
the LC system at 7.1% Pump B.
–– Alternatively, prepare manually by mixing 710 μl of His60 Ni Elution Buffer with 9.29 ml of His60 Ni
Equilibration Buffer.
Elution Buffer: 50 mM sodium phosphate, 300 mM sodium chloride, 300 mM imidazole; pH 7.4
Regeneration Buffer: 20 mM MES (2-(N-morpholine)-ethanesulfonic acid), 0.3 M sodium chloride; pH
5.0
Imidazole: Use a highly pure, low-absorbance imidazole ideal for LC applications
(Fisher, Product No. BP 305-50).
PT5143-1
031716
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Clontech Laboratories, Inc. A Takara Bio Company
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