Product Overview Data Sheet Protocol
pFB-CHIg-mG2ae1 is a cloning vector that expresses the mouse IgG2a heavy chain constant region with L235E/E318A/K320A/K322A mutations. It is a constitutive mammalian expression vector designed to deliver exceptionally high levels of antibody expression. This circular vector features an enhanced, full-length CMV promoter and other expression elements that typically enable higher expression levels. It can be used in suspension-adapted cells, such as Expi293F™ and ExpiCHO™, for transient protein expression. Additionally, it can serve as a Geneticin®-selectable expression plasmid for engineering stable cell lines. The vector carries an ampicillin resistance gene.
Characteristics
Fc engineered mouse IgG2a expression with L235E/E318A/K320A/K322A mutation:
– Reduced binding to FcγRI and C1q
– Reduced ADCC and CDC
| Antibiotic Resistance | Ampicillin (AmpR) |
| Constitutive or Inducible System | Constitutive |
| Delivery Type | Transfection |
| Promoter | CMV |
| Product Type | Mammalian Expression Vector |
| Cloning Method | Restriction Enzyme (5’-AgeI; 3’-XhoI) or Homologous Assembly |
Contents & Storage
- 20 µg of pFB-CHIg-mG2ae1 in Tris-EDTA buffer
- Store at -20°. Vectors are guaranteed stable for 6 months when properly stored.
Materials required for Fc engineered antibody generation
- pFB-CLIg-mk or pFB-CLIg-ml1 or pFB-CLIg-ml2 plasmid expressing the mouse kappa or lambda light chain.
Steps for Fc engineered antibody generation
- Cloning your heavy chain variable region (VH) into pFB-CHIg-mG2ae1 vector to make mouse heavy chain expression plasmid.
- Cloning your light chain variable region (VL) into pFB-CLIg-mk or pFB-CLIg-ml1 or pFB-CLIg-ml2 vector to make mouse light chain expression plasmid.
- Co-transfecting both heavy chain and light chain expression plasmids into your desired mammalian cell (such as CHO, HEK293) for Fc engineered antibody production.
References
- Duncan et al., 1988. The binding site for C1q on IgG. Nature 332, 738–740.
- Wines et al., 2000. The IgG Fc contains distinct Fc receptor (FcR) binding sites: the leukocyte receptors Fc gamma RI and Fc gamma RIIa bind to a region in the Fc distinct from that recognized by neonatal FcR and protein A. J Immunol 164, 5313–5318.
