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Human FAS / CD95 / APO-1 / TNFRSF6 HEK293 Cell Lysate (WB positive control)

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Human CD95 Transfected / Overexpression Cell Lysate Product Information
Expressed Host:Human Cells
Product Description:Human Cell lysate that Human CD95 / Fas / TNFRSF6 transfected / overexpressed for Western blot (WB) positive control. The whole cell lysate is provided in 1X Sample Buffer (1X modified RIPA buffer+1X SDS loading buffer).
Sequence information:A DNA sequence encoding the extracellular domain (Met 1-Asn 173) of human FAS antigen (NP_000034.1) was expressed, with a C-terminal polyhistidine tag.
Predicted N Terminal:Gln 26
Molecule Mass:The secreted recombinant human FAS comprises 159 amino acids and has a predicted molecular mass of 18.2 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of rhFAS is approximately 30-40 kDa due to glycosylation.
Human CD95 Transfected / Overexpression Cell Lysate Usage Guide
Preparation Method:Cell lysate was prepared by homogenization 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.
Lysis Buffer: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.
Quality Control Testing:12.5% SDS-PAGE Stained with Coomassie Blue after protein purification.
Stability:Samples are stable for up to twelve months from date of receipt.
Recommend Usage: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. 3. Store the lyophilized cell lysate at 4℃. After re-dissolution, recommend to aliquot it into smaller quantities and store at -80℃.
Storage Buffer:1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Storage Instruction:Store at 4℃. After re-dissolution, aliquot and store at -80℃.
Application notes:Western blot (WB): Use at an assay dependent dilution.
Other Applications: Not tested.
Optimal dilutions/concentrations should be determined by the end user.
CD95/APO-1/TNFRSF6 Background

CD95 (APO-1/Fas) is an important inducer of the extrinsic apoptosis signaling pathway and therapy induced apoptosis of many tumor cells has been linked to the activity of CD95. is a prototype death receptor characterized by the presence of an 80 amino acid death domain in its cytoplasmic tail. This domain is essential for the recruitment of a number of signaling components upon activation by either agonistic anti-CD95 antibodies or cognate CD95 ligand that initiate apoptosis. The complex of proteins that forms upon triggering of CD95 is called the death-inducting signaling complex (DISC). The DISC consists of an adaptor protein and initiator caspases and is essential for induction of apoptosis. CD95 is also crucial for the negative selection of B cells within the germinal center (GC). Impairment of CD95-mediated apoptosis results in defective affinity maturation and the persistence of autoreactive B-cell clones. Changes in the expression of CD95 and/or its ligand CD95L are frequently found in human cancer. The downregulation or mutation of CD95 has been proposed as a mechanism by which cancer cells avoid destruction by the immune system through reduced apoptosis sensitivity. Thus, CD95 has therefore been viewed as a tumor suppressor. CD95 has been reported to be involved in the activation of NF-kappaB, MAPK3/ERK1, MAPK8/JNK, and the alternate pathways for CTL-mediated cytotoxicity. Accordingly, this protein is implicated in the pathogenesis of various malignancies and diseases of the immune system. The CD95/CD95L system was implicated in the etiology of inflammatory bowel disease (IBD) based, primarily, on the finding that CD95 is highly expressed in the intestinal epithelial cells and that epithelial apoptosis is increased in IBD.

Human CD95/APO-1/TNFRSF6 References
  • Mschen M, et al. (2002) The origin of CD95-gene mutations in B-cell lymphoma. Trends Immunol. 23(2): 75-80.
  • Peter ME, et al. (2003) The CD95(APO-1/Fas) DISC and beyond. Cell Death Differ. 10(1): 26-35.
  • Peter ME, et al. (2005) Does CD95 have tumor promoting activities Biochim Biophys Acta. 1755(1): 25-36.
  • Chen L, et al. (2010) Cell death in the colonic epithelium during inflammatory bowel diseases: CD95/Fas and beyond. Inflamm Bowel Dis. 16(6): 1071-6.
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    Catalog: 10217-H08HL-300
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