产品名称：HistaggedProteinPurificationKit
品牌：Engibody
货号：P3406
关键词：His蛋白纯化试剂盒
Description
His-TaggedProteinPurificationKit(Ni-NTAAgaroseResin6FF,SolubleProtein)
Components
onecolumn;4mlNi2+Sepharose4B;5gNiSO4;6gimidazole;10gEDTA
Storageinstruction
Storeat+4°Canddonotfreeze.
Protocols
Procedure
1).ResuspendtheNi2+Sepharose4Bbytappingandinvertingthevialseveraltimesimmediatelybeforedispensing.Donotvortex.Thentransfer2mlintothecolumn,thegelwillsedimentatenaturally.
2).Washthecolumnwith5bedvolumesofwater,thenequilibratethecolumnwith5bedvolumesofbindingbuffer（20mmol/Lphosphatebuffer,500mmol/LNaCl,pH7.8）.
3).Centrifugecelllysate（20mmol/Lphosphatebuffer,500mmol/LNaCl,pH7.8）at12,000g,collectthesupernatantandloaditintothecolumnatarateof0.1mlpermin.
4).Washthecolumnwith5-10bedvolumesofBindingbufferatarateof0.1mlpermin.
5).EluteSeparatelywithElutionbufferofimidazolesolutionsatdiferrentconcentration(usually10,20,50,100,150,200mmol/L,dependingonhistaggedfusionprotein).Thencollecteveryeluate.
6).SDS-PAGEgelelectrophoresiscananalyzethepurityofhistaggedfusionprotein).
Regeneration
1).Washthecolumnwith2bedvolumesof0.2mol/LAceticacid,thenWashthecolumnwith5bedvolumesofwater.
2).Washthecolumnwith5bedvolumesof50％ethanol,thenWashthecolumnwith5bedvolumesofwater.
3).Washthecolumnwith5bedvolumesof0.1mol/LEDTA,thenWashthecolumnwith5bedvolumesofwater.
4).Washthecolumnwith2bedvolumesof0.1mol/LNiSO4,thenWashthecolumnwith5bedvolumesofwater.
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