This Human IFI30 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of IFI30 protein (Cat: 12623-H08H) from the overexpression lysate was verified.
A DNA sequence encoding the human IFI30 (P13284) (Met1-Lys243) with a C-terminal polyhistidine tag was expressed.
The recombinant human IFI30 is a disulfide-linked homodimer. The reduced monomer comprises 217 amino acids and has a predicted molecular mass of 24.7 kDa. The apparent molecular mass of the protein is approximately 31-34 kDa in SDS-PAGE under reducing conditions.
Human IFI30 HEK293 Overexpression Lysate: 使用指南
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube.
2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
稳定性 & 储存条件
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Western Blot (WB) Optimal dilutions/concentrations should be determined by the end user.
Human IFI30 HEK293 Overexpression Lysate: 别称
Human GILT Overexpression Lysate; Human IFI-30 Overexpression Lysate; Human IFI30 Overexpression Lysate; Human IP-30 Overexpression Lysate; Human IP30 Overexpression Lysate
IFI30 belongs to the GILT family. This family includes the two characterized human gamma-interferon-inducible lysosomal thiol reductase (GILT) sequences: P13284 and Q9UL08. It also contains several other eukaryotic putative proteins with similarity to GILT. The aligned region contains three conserved cysteine residues. Besides, the two GILT sequences possess a C-X(2)-C motif that is shared by some of the other sequences in the family. This motif is thought to be associated with disulfide bond reduction. IFI30 is a lysosomal thiol reductase that can reduce protein disulfide bonds. It facilitates the generation of MHC class II-restricted epitopes from disulfide bond-containing antigens by the endocytic reduction of disulfide bonds. It also facilitates MHC class I-restricted recognition of exogenous antigens containing disulfide bonds by CD8+ T-cells or cross-presentation. IFI30 may facilitate the complete unfolding of proteins destined for lysosomal degradation and plays an important role in antigen processing.
interferon gamma inducible protein 30
Haque MA, et al. (2002) Absence of gamma-Interferon-inducible Lysosomal Thiol Reductase in Melanomas Disrupts T Cell Recognition of Select Immunodominant Epitopes. J Exp Med. 195(10):1267-77.
Phan UT, et al. (2000) Gamma-interferon-inducible lysosomal thiol reductase (GILT). Maturation, activity, and mechanism of action. J Biol Chem. 275(34):25907-14.
Phan UT, et al. (2001) Multiple species express thiol oxidoreductases related to GILT. Immunogenetics. 53(4):342-6.