RecombiMAb human IgG4 (S228P/R409K) isotype control, anti-hen egg lysozyme

Catalog #CP183

$224.00 - $6,087.00

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Product Details

This human IgG4 S228P/R409K isotype control antibody reacts with hen egg lysozyme and has low or no specific binding to any human sample.Ā  The S228P mutation is included to prevent IgG4 Fab exchange. The addition of the R409K mutation has been shown to prevent acid-induced aggregation of human IgG4. This recombinant human IgG4 antibody is produced in CHO cells.

Specifications

Isotype Human IgG4, Īŗ
Recommended Dilution Buffer InVivoPure pH 7.0 Dilution Buffer
Mutations S228P/R409K
Immunogen Hen egg lysozyme (HEL)
Formulation PBS, pH 7.0
Contains no stabilizers or preservatives
Endotoxin <1EU/mg (<0.001EU/Ī¼g)
Determined by LAL gel clotting assay
Purity >95%
Determined by SDS-PAGE
Sterility 0.2 Āµm filtration
Production Purified from CHO cell supernatant in an animal-free facility
Purification Protein A
Molecular Weight 150 kDa
Murine Pathogen Tests Ectromelia/Mousepox Virus: Negative
Hantavirus: Negative
K Virus: Negative
Lactate Dehydrogenase-Elevating Virus: Negative
Lymphocytic Choriomeningitis virus: Negative
Mouse Adenovirus: Negative
Mouse Cytomegalovirus: Negative
Mouse Hepatitis Virus: Negative
Mouse Minute Virus: Negative
Mouse Norovirus: Negative
Mouse Parvovirus: Negative
Mouse Rotavirus: Negative
Mycoplasma Pulmonis: Negative
Pneumonia Virus of Mice: Negative
Polyoma Virus: Negative
Reovirus Screen: Negative
Sendai Virus: Negative
Theilerā€™s Murine Encephalomyelitis: Negative
Storage The antibody solution should be stored at the stock concentration at 4Ā°C. Do not freeze.
Namisaki H, Saito S, Hiraishi K, Haba T, Tanaka Y, Yoshida H, Iida S, Takahashi N. (2020). "R409K mutation prevents acid-induced aggregation of human IgG4" PLoS One 15(3):e0229027. PubMed

Human immunoglobulin G isotype 4 (IgG4) antibodies are suitable for use in either the antagonist or agonist format because their low effector functions prevent target cytotoxicity or unwanted cytokine secretion. However, while manufacturing therapeutic antibodies, they are exposed to low pH during purification, and IgG4 is more susceptible to low-pH-induced aggregation than IgG1. Therefore, we investigated the underlying mechanisms of IgG4 aggregation at low pH and engineered an IgG4 with enhanced stability. By swapping the constant regions of IgG1 and IgG4, we determined that the constant heavy chain (CH3) domain is critical for aggregate formation, but a core-hinge-stabilizing S228P mutation in IgG4 is insufficient for preventing aggregation. To identify the aggregation-prone amino acid, we substituted the CH3 domain of IgG4 with that of IgG1, changing IgG4 Arg409 to a Lys, thereby preventing the aggregation of the IgG4 variant as effectively as in IgG1. A stabilizing effect was also recorded with other variable-region variants. Analysis of thermal stability using differential scanning calorimetry revealed that the R409K substitution increased the Tm value of CH3, suggesting that the R409K mutation contributed to the structural strengthening of the CH3-CH3 interaction. The R409K mutation did not influence the binding to antigens/human FcĪ³ receptors; whereas, the concurrent S228P and R409K mutations in IgG4 suppressed Fab-arm exchange drastically and as effectively as in IgG1, in both in vitro and in vivo in mice models. Our findings suggest that the IgG4 R409K variant represents a potential therapeutic IgG for use in low-effector-activity format that exhibits increased stability.