Human A33 HEK293 Overexpression Lysate

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Human A33 HEK293 Overexpression Lysate: 产品信息

产品描述
This Human A33 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of A33 protein (Cat: 11277-H08H) 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 extracellular domain of human GPA33 (Q99795-1) (Met 1-Val 235) was fused with a polyhistidine tag at the C-terminus.
分子量
The secreted recombinant human GPA33 consists of 225 amino acids and has a predicted molecular mass of 25 kDa. The apparent molecular mass of the rhGPA33 is approximately 35-40 kDa in SDS-PAGE under reducing conditions due to glycosylation.

Human A33 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 A33 HEK293 Overexpression Lysate: 别称

Human A33 Overexpression Lysate

A33 背景信息

Cell surface A33 antigen, also known as glycoprotein A33, is a single-pass type I membrane protein that is expressed in the normal gastrointestinal epithelium and 95% of colon cancers. GPA33 contains one Ig-like C2-type (immunoglobulin-like) domain and one Ig-like V-type (immunoglobulin-like) domain. The open reading frame encodes a 319-amino acid polypeptide having a putative secretory signal sequence and 3 potential glycosylation sites. The predicted mature protein has a 213-amino acid extracellular region, a single transmembrane domain, and a 62-amino acid intracellular tail. Intracellular traffic and recycling to the cell surface appear to play a major role in GPA33 function and to have an influence on its surface density superseding translational regulation. GPA33 has become a promising target of immunologic therapy strategies, but its biologic function and potential role in tumorigenesis are unknown. EpCAM protein and GPA33 mRNA expressions are specific and sensitive markers of Barrett's metaplasia (BM). GPA33 may also play a role in cell-cell recognition and signaling.
全称
glycoprotein A33 (transmembrane)
研究领域
参考文献
  • Heath J.K., et al., 1997, Proc. Natl. Acad. Sci. USA. 94:469-74.
  • Ritter G., et al., 1997, Biochem. Biophys. Res. Commun. 236:682-6.
  • Frey,D. et al., 2008, Cancer Biother Radiopharm  23 (1):65-73.
  • Rageul,J. et al., 2009, Int J Cancer 125 (12):2802-9. 
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