Human FCGR3A / CD16a (V176) Protein

INTRODUCTION

QC & Assay Protocol

Citation & Background

Synonym
FCGR3A,CD16A,FCG3,FCGR3,IGFR3
Source
Human FCGR3A / CD16a (V176) Protein,also called Human FCGR3A / CD16a Protein (176 Val), Gly 17 - Gln 208 (Accession # P08637-1), was produced in human 293 cells (HEK293) at ACROBiosystems.
Predicted N-terminus: Gly 17
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Molecular Characterization
This protein carries a polyhistidine tag at the C-terminus.
The protein has a calculated MW of 23.7 kDa. The protein migrates as 40-50 kDa under reducing (R) condition (SDS-PAGE) due to glycosylation.
Endotoxin
Less than 1.0 EU per μg by the LAL method.
Purity
>95% as determined by reduced SDS-PAGE.

Formulation
Lyophilized from 0.22 μm filtered solution in PBS, pH7.4. Normally trehalose is added as protectant before lyophilization.
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Reconstitution
Please see Certificate of Analysis for specific instructions.
For best performance, we strongly recommend you to follow the reconstitution protocol provided in the CoA.
Storage
For long term storage, the product should be stored at lyophilized state at -20°C or lower.
Please avoid repeated freeze-thaw cycles.
No activity loss was observed after storage at:
1. 4-8^°C for 12 months in lyophilized state;
2. -70^°C for 3 months under sterile conditions after reconstitution.

SDS-PAGE

Human FCGR3A / CD16a (V176), His Tag on SDS-PAGE under reducing (R) condition. The gel was stained overnight with Coomassie Blue. The purity of the protein is greater than 95%.

Bioactivity-ELISA

Human FCGR3A / CD16a (V176), His TagHuman FCGR3A / CD16a (V176), His Tag (Cat. No. CD8-H52H4) ELISA bioactivity

Immobilized Yervoy Ipilimumab (Human IgG1) at 5 μg/mL (100 μL/well) can bind Human FCGR3A / CD16a (V176), His Tag (Cat. No. CD8-H52H4 ) with a linear range of 1-16 ng/mL (QC tested).

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Bioactivity-SPR

Immobilized Human CD32a (V167) Protein (Cat. No. CD8-H52H4) on CM5 Chip via anti-His antibody, can bind Herceptin with an affinity constant of 163 nM as determined in SPR assay (Biacore T200) (QC tested).

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Citations
- Charge variant analysis of proposed biosimilar to Trastuzumab
  Authors: Dakshinamurthy P, et al.
  Journal: Biologicals 2017
  Application: SPR
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Background
CD16 is a low affinity Fc receptor, and has been identified as Fc receptors FcγRIIIa (CD16a) and FcγRIIIb (CD16b). These receptors bind to the Fc portion of IgG antibodies. CD16 encoded by two different highly homologous genes in a cell type-specific manner.CD16 is found on the surface of natural killer cells, neutrophil polymorphonuclear leukocytes, monocytes and macrophages. CD16a antigen is also known as Low affinity immunoglobulin gamma Fc region receptor III-A, Fc-gamma RIII-alpha. CD16b is a low-affinity, GPI-linked receptor expressed by neutrophils and eosinophils, whereas CD16a is an intermediate affinity polypeptide-anchored transmembrane glycoprotein expressed natural killer cells, macrophages, subpopulation of T-cells, immature thymocytes and placentaltrophoblasts.CD16a is involved in phagocytosis, secretion of enzymes and inflammatory mediators, antibodydependent cytotoxicity and clearance of immune complexes. Aberrant expression or mutations of CD16a is implicated in susceptibility to recurrent viral infections, systemic lupus erythematosus, and alloimmune neonatal neutropenia.
References
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