>Fc Receptor Proteins
The efficacy of a therapeutic antibody not only depends on the Fab fragment and its binding activity to the target antigen, but also depends on the Fc fragment and its interaction with key Fc receptors. The binding affinity of the Fc fragment towards FcRn (FCGRT&B2M) would predict the antibody’s half-life, while that between the Fc fragment and FcγRIIIa (CD16a) would influence the antibody’s ability to elicit ADCC (antibody dependent cell mediated cytotoxicity). Therefore, candidates must be tested against a panel of receptors during antibody engineering.
ACROBiosystems offers a comprehensive collection of recombinant Fc receptor proteins, including their common variants, to help expedite your mAb development.
To meet the high purity requirement of pharmaceutical applications, our production team perform both SDS-PAGE and HPLC analyses to the Fc receptors for QC purposes. It’s noteworthy that it’s essential to use monomeric Fc receptor, which represents its natural state. The formation of oligomers during production may result in artificially enhanced binding interaction due to avidity effect. To avoid such situation, we have established strict internal control standards of monomer purity by HPLC testing. Only those batches meeting all purity requirements are released.
Fig. 1 The purity of Human Fc gamma RIIA / CD32a (R167) Protein (Cat. No. CDA-H5221) is greater than 95% as determined in a HPLC analysis.
The binding affinity between two molecules can be determined by many methods. However, the studies of Fc interaction are often done by SPR, because traditional ELISA is not appropriate for low affinity receptors such as CD16 and CD32. To assure our Fc receptors deliver expected performance, our QC team use Biacore platform to test our products against reference antibodies. Additionally, to meet more applications in the development of antibody drugs, our QC team also use ForteBio platform to test our products against reference antibodies. All SPR&BLI protocols are available for free.
Fig. 2 Human CD64, His Tag (SPR & BLI verified) (Cat. No. FCA-H52H1) captured on CM5 chip via anti-His antibody can bind Herceptin with an affinity constant of 4.92 nM as determined in a SPR assay (Biacore 8K) (QC tested).
Fig. 3 Immobilized Human FCGRT&B2M Heterodimer Protein, His Tag&Strep II Tag (SPR & BLI verified) (Cat. No. FCM-H5286) on CM5 Chip via Anti-His antibody, can bind Herceptin with an affinity constant of 0.261 μM as determined in a SPR assay (Biacore T200).
Fig. 4 Loaded Human CD64, His Tag (SPR & BLI verified) (Cat. No. FCA-H52H1) on HIS1K Biosensor, can bind Herceptin with an affinity constant of 6.98 nM as determined in BLI assay (ForteBio Octet Red96e).
Fig. 5 Loaded Human FCGRT&B2M Heterodimer Protein, His Tag (SPR & BLI verified) (Cat. No. FCN-H52W7) on SA Biosensor via Biotin his antibody, can bind Herceptin with an affinity constant of 0.14 μM as determined in BLI assay (ForteBio Octet Red96e).
We routinely apply rigorous quality control measures to ensure consistent performance of our product. As shown below, the batch variation among the tested samples is negligible.
Fig. 6 Immobilized Human Fc gamma RIIB / CD32b Protein (Cat. No. CDB-H5228) on CM5 Chip via anti-His antibody, can bind Rituximab with an affinity constant of 10 μM as determined in a SPR assay (Biacore T200).
We have compared the performance of different batches of CD32b (CDB-H5228) in the same assay, and the software analysis showed that the similarity score was very high, meaning high batch-to-batch consistency.
Fig. 7 Batch consistency of Human Fc gamma RIIB / CD32b (Cat. No. CDB-H5228). The consistency of different batches is more than 90%.
The use of biotin labeling can make your assay development much easier. We offer a variety of ready-to-use biotinylated Fc receptors. These proteins are produced using our in-house developed labeling techniques, which confers high bioactivity and minimal batch-to-batch variation. In addition, we have developed a series of Fc receptors of other species, which are suitable for the screening of non-humanized antibodies or the species cross reaction.
Fig. 8 Biotinylated Human CD16a (V176), His Tag, Avi Tag (Cat. No.CDA-H82E9) captured on Biotin CAP- Series S Sensor Chip can bind Rituximab with an affinity constant of 0.261 μM as determined in a SPR assay (Biacore T200).
Fig. 9 Immobilized Cynomolgus / Rhesus macaque FcRn Protein (Cat. No. FCM-C5284) on CM5 Chip via anti-His antibody, can bind Herceptin with an affinity constant of 0.403 μM as determined in a SPR assay (Biacore T200).
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