Human Alkaline phosphatase HEK293 Overexpression Lysate

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Human Alkaline phosphatase HEK293 Overexpression Lysate: 产品信息

产品描述
This Human Alkaline phosphatase overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of Alkaline phosphatase protein (Cat: 13225-H02H) from the overexpression lysate was verified.
表达宿主
HEK293 Cells
种属
Human
蛋白构建信息
A DNA sequence encoding the mature form of human ALPI (P09923) (Met 1-Asp 503),without the pro peptide, was fused with a polyhistidine tag at the C-terminus.
分子量
The secreted recombinant human ALPI/Fc is a disulfide-linked homodimeric protein. The reduced monomer consists of 725 amino acids and has a predicted molecular mass of 79.5 kDa. The apparent molecular mass of rhALPI /Fc monomer is approximately 90-95 kDa in SDS-PAGE under reducing conditions due to glycosylation.

Human Alkaline phosphatase HEK293 Overexpression Lysate: 使用指南

制备方法
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.
裂解缓冲液
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
使用建议
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.
缓冲液
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.
应用
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human Alkaline phosphatase HEK293 Overexpression Lysate: 别称

Human IAP Overexpression Lysate

Alkaline phosphatase 背景信息

ALPI encodes for intestinal phosphatase alkaline, a brush border metalloenzyme that hydrolyses phosphate from the lipid A moiety of lipopolysaccharides and thereby drastically reduces Toll-like receptor 4 agonist activity. ALPI mutations impaired either stability or catalytic activity of ALPI and rendered it unable to detoxify lipopolysaccharide-dependent signalling. ALPI mutations should be included in screening for monogenic causes of inflammatory bowel diseases and lay the groundwork for ALPI-based treatments in intestinal inflammatory disorders.
全称
alkaline phosphatase, intestinal
参考文献
  • Nakano T, et al. (2009) Role of lysophosphatidylcholine in brush-border intestinal alkaline phosphatase release and restoration. Am J Physiol Gastrointest Liver Physiol. 297(1): G207-14.
  • Henthorn PS, et al. (1987) Nucleotide and amino acid sequences of human intestinal alkaline phosphatase: close homology to placental alkaline phosphatase. Proc Natl Acad Sci. 84(5): 1234-8.
  • Oetting WS, et al. (2003) Oculocutaneous albinism type 1: the last 100 years. Pigment Cell Res. 16(3): 307-11.
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