Human CD16a HEK293 Overexpression Lysate

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Human CD16a HEK293 Overexpression Lysate: Product Information

Product Description
This Human CD16a overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of CD16a protein (Cat: 10389-H27H) from the overexpression lysate was verified.
Expression Host
HEK293 Cells
Species
Human
Sequence Information
A DNA sequence encoding the extracellular domain (Met 1-Gln 208) of human CD16a (P08637-1) was fused with a c-terminal polyhistidine tagged AVI tag at the C-terminus. It is identical to FCGR3A158F/V in the reference.
Molecule Mass
The secreted recombinant human CD16a consists of 223 amino acids and predicts a molecular mass of 25.1 kDa. By SDS-PAGE under reducing conditions, the apparent molecular mass of the protein is approximately 46 kDa due to glycosylation.

Human CD16a HEK293 Overexpression Lysate: Usage Guide

Preparation Method
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.
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.
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.
Sample Buffer
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.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human CD16a HEK293 Overexpression Lysate: Alternative Names

Human CD16 Overexpression Lysate; Human CD16A Overexpression Lysate; Human Fc gamma RIIIa Overexpression Lysate; Human FCG3 Overexpression Lysate; Human FCGR3 Overexpression Lysate; Human FCGRIII Overexpression Lysate; Human FCR-10 Overexpression Lysate; Human FCRIII Overexpression Lysate; Human FCRIIIA Overexpression Lysate; Human IGFR3 Overexpression Lysate; Human IMD20 Overexpression Lysate

CD16a Background Information

The Fc receptor with low affinity for IgG (FCGR3, or CD16) is encoded by 2 nearly identical genes, FCGR3A and FCGR3B, resulting in tissue-specific expression of alternative membrane-anchored isoforms. FCGR3A, it is also known as CD16a, encodes a transmembrane protein expressed on activated monocytes/macrophages, natural killer (NK) cells, and a subset of T cells. CD16a / FCGR3A is a receptor expressed on NK cells that facilitates antibody dependent cellular cytotoxicity (ADCC) by binding to the Fc portion of various antibodies. CD16a / FCGR3A also has a broader function. CD16a / FCGR3A is directly involved in the lysis of some virus-infected cells and tumor cells by NK cells, independent of antibody binding. Cross-linking of CD16a / FCGR3A on NK cells resulted in increased intracellular Ca2+ levels and a cascade of biochemical events similar to those activated by the T cell receptor. CD16a / FCGR3A on human NK cells is a lysis receptor that mediates the direct killing of some virus infected and tumor cells, independent of antibody ligation.
Full Name
Fc fragment of IgG, low affinity IIIa, receptor (CD16a)
References
  • David Dornan, et al. Effect of FCGR2A and FCGR3A variants on CLL outcome. Blood. 2010 116: 4212-4222
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