Human APCDD1 HEK293 Overexpression Lysate: Product Information
This Human APCDD1 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of APCDD1 protein (Cat: 15383-H02H) from the overexpression lysate was verified.
A DNA sequence encoding the human APCDD1 (NP_694545.1) (Met1-Gly486) was expressed with the Fc region of human IgG1 at the C-terminus.
The recombinant human APCDD1/Fc comprises 701 amino acids and has a predicted molecular mass 79.6 kDa. The apparent molecular mass of the protein is approximately 85.7 kDa in SDS-PAGE under reducing conditions.
Human APCDD1 HEK293 Overexpression Lysate: Usage Guide
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).
Stability & Storage
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 APCDD1 HEK293 Overexpression Lysate: Alternative Names
Human B7323 Overexpression Lysate; Human DRAPC1 Overexpression Lysate; Human FP7019 Overexpression Lysate; Human HHS Overexpression Lysate; Human HTS Overexpression Lysate; Human HYPT1 Overexpression Lysate
APCDD1 Background Information
Osteosarcoma (OS) is the most common type of bone tumor in children and adults. The expression of APCDD1, a Wnt antagonist, was reduced in OS tissues and cells compared to adjacent normal tissue and osteoblast cells, respectively. Mechanistically, this was due to increased levels of methylation in the promoter region of the APCDD1 gene. Consistently, the DNA methyltransferase inhibitor 5-AZA-dC, reduced DNA methylation in the APCDD1 promoter, and restored APCDD1 expression in OS tissue and cells. Moreover, DNMT3a, but not DNMT1 or DNMT3b, was the major DNA methyltransferase that facilitated hyper-methylation of DNA in the APCDD1 promoter, thus reducing APCDD1 mRNA levels in OS tissues. Importantly, ectopic expression of APCDD1 suppressed activity of the Wnt/beta-Catenin signaling pathway in OS cells and inhibited their invasion and reversed their EMT-like properties, while depletion of APCDD1 promoted invasion and metastasis of osteosarcoma cells in vitro and in vivo. That APCDD1 modulates the gene expression of Wnt- and EK-related signaling molecules at the cap stage of tooth development, and is involved in tooth cusp patterning by modulating the epithelial rearrangement in the IEE. In hair follicle cells APCDD1 inhibits the canonical WNT/beta-Catenin pathway and its inactivation is associated with an autosomal dominant form of hair loss. APCDD1 sustains the expression and activation of beta-Catenin.
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