CARD8 Overexpression Lysate

Western Blot: CARD8 Overexpression Lysate (Adult Normal) [NBP2-10259] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for CARD8.

• Reactivity: Hu
• Applications: WB

CARD8 Overexpression Lysate Summary

• Description:

  CARD8 Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC230245
  
  Accession#: NM_001184900
  
  Protein Tag: C-MYC/DDK
  
  You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.
• Gene: CARD8

Applications/Dilutions

• Dilutions:
  • Western Blot
• Application Notes: This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (NBP1-71705) present on the protein construct.
  
  Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.

Packaging, Storage & Formulations

• Storage: Store at -80C. Avoid freeze-thaw cycles.
• Buffer: RIPA buffer

Lysate Details for Array

• Type: Overexpression

Notes

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

Alternate Names for CARD8 Overexpression Lysate

• Apoptotic protein NDPP1
• CARD inhibitor of NF-kappaB-activating ligands
• CARDINALFLJ18121
• CARD-inhibitor of NF-kappa-B-activating ligand
• caspase recruitment domain family, member 8
• caspase recruitment domain-containing protein 8
• DACAR
• DAKAR
• KIAA0955DKFZp779L0366
• NDPP
• NDPP1MGC57162
• TUCANFLJ18119
• Tumor up-regulated CARD-containing antagonist of CASP9
• tumor up-regulated CARD-containing antagonist of caspase nine

Background

TUCAN (also known as CARD8 and CARDINAL) is a CARD domain containing protein. TUCAN stands for tumor-up-regulated CARD-containing antagonist of caspase nine. Proteins containing a CARD (caspase-associated recruitment domain) domain are key regulators of cell death, cell survival and cytokine production (reviewed in Damiano and Reed, 2004). CARD domains are found in the N-terminal pro-domains of certain caspases, a family of apoptotic and pro-inflammatory proteases, as well as in a diversity of other proteins including TUCAN. CARD domains are homotypic protein interaction motifs that enable networks of proteins to communicate via CARD-CARD interactions. TUCAN is an anti-apoptotic CARD protein that can protect tumors from cell death stimuli, and is overexpressed in certain forms of cancer. The role of TUCAN in tumor biology remains to be fully elucidated, and there appears to be be various mechanisms by TUCAN can potentially contribute to apoptosis resistance (reviewed in Checinska et al, 2006). For example, TUCAN has been shown to inhibit caspase-9 activation by binding to the CARD region of pro-caspase-9, thereby suppressing the formation of the Apaf-1-caspase-9 apoptotic complex and apoptosis. Additionally, a TUCAN isoform has been described that blocks both caspase-8 and caspase-9 mediated apoptosis. However, in some tumors, TUCAN does not appear to interact with caspase-9 and may play a role in modulating NF-kB transcription factor survivial signaling pathways. In this regard a CARD-independent interaction of TUCAN with Ikkg has been described, resulting in the inhibition of interleukin-1 and TNF-induced NF-kB activation. Human TUCAN is a 431 amino acid protein according to GenBank no. gi|14424229|sp|Q9Y2G2|CARD8_HUMAN, and migrates at ~45-49 kDa on SDS-PAGE.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

Bioinformatics

• Gene Symbol: CARD8