Anti-Natalizumab Antibodies

Antibodies for bioanalytical ligand binding assays to measure natalizumab and biosimilar products

Ready-made recombinant monoclonal anti-idiotypic antibodies, highly specific for the humanized monoclonal antibody drug natalizumab (Tysabri).

Anti-Natalizumab Inhibitory Antibodies

These antibodies inhibit the binding of the humanized monoclonal antibody drug natalizumab to its target, human alpha4-beta1-integrin, and therefore detect free drug. An antibody pair is ideal for the development of a pharmacokinetic (PK) bridging ELISA. Two antibodies of high and medium affinity are suitable as a reference standard for an anti-drug antibody (ADA) assay.

Our recombinant monoclonal antibodies are generated using Human Combinatorial Antibody Library (HuCAL®) technology and CysDisplay®, a proprietary method of phage display, and employing guided selection methods to obtain highly targeted reagents. The result is highly specific and high affinity antibodies, ideal for development of PK assays. HuCAL antibodies are fully human, and when in full length IgG format they can be used as a control or calibrator in an ADA assay. The in vitro production of recombinant antibodies means that there is a consistent, secure supply throughout preclinical development and clinical trials, avoiding the need to develop new reagents.

More information about anti-idiotypic antibodies and their binding types and properties

Anti-biotherapeutic antibody quality control and characterization

Table 1. Antibodies Specific to Natalizumab.

1 F=DYKDDDDK-tag; H=His-6-tag
2 Affinity measured in the monovalent Fab format

Related Products

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Natalizumab biosimilar for research use Human Anti-Integrin Alpha 4 Antibody (MCA6104)

Pharmacokinetic Assay – Bridging ELISA Format

Schematic image of PK Bridging ELISA. Anti-idiotypic capture antibody, Fab format (purple), monoclonal antibody drug (gold), anti-idiotypic detection antibody, Ig format (blue), labeled with HRP.

Fig. 1. Natalizumab PK assay bridging format using antibodies HCA248 and HCA249P.

Measurement of free drug with human anti-natalizumab antibodies

In figure 1, a microtiter plate was coated overnight with Anti-Natalizumab Antibody, clone AbD21375 (HCA248) at a concentration of 1 µg/ml. After washing and blocking with PBST + 5% BSA, 10% human serum was added spiked with increasing concentrations of natalizumab. Detection was performed using HRP conjugated Anti-Natalizumab Antibody, clone AbD21375_hIgG1 (HCA249P) at a concentration of 2 µg/ml in HISPEC Assay Diluent (BUF049A) and QuantaBlu Fluorogenic Peroxidase Substrate. Data are shown as the mean of three measurements.

Protocol: Natalizumab PK bridging ELISA using antibodies HCA248 and HCA249P

Inhibition ELISA

Fig. 2. Inhibition of natalizumab binding to human alpha4-beta1-integrin by HCA248.

In figure 2, a microtiter plate was coated overnight with human alpha4-beta1-integrin at a concentration of 1 µg/ml. After washing and blocking with PBST + 5% BSA, a pre-incubated mixture of natalizumab (0.3 µg/ml) plus increasing concentrations of Human Anti-Natalizumab Antibody, clone AbD21375 (HCA248) in the monovalent Fab format was added. Free natalizumab, still capable of binding to the human alpha4-beta1-integrin coated plate, was detected using Mouse Anti Human IgG (Fc) CH2 Domain:HRP Antibody (MCA647P) and QuantaBlu Fluorogenic Peroxidase Substrate. Data are shown as the mean of three measurements.

Anti-Drug Antibody Assay - Bridging ELISA Format

Two anti-natalizumab antibodies are available in fully human IgG1 format, each with a different affinity for the target, allowing the ADA assay to be tailored to the specific needs of the developer.

Schematic image of ADA bridging assay. Monoclonal antibody drug as capture antibody and detection antibody labeled with HRP (gold), fully human anti-idiotypic antibody, Ig format (blue).

Fig. 3. ADA bridging ELISA with HCA249 or HCA250.

In figure 3, a microtiter plate was coated overnight with natalizumab at a concentration of 1 µg/ml. After washing and blocking with PBST + 5% BSA, PBST with 10% human serum was added spiked with increasing concentrations of human Anti-Natalizumab Antibody, either HCA249 or HCA250. Detection was performed using HRP conjugated natalizumab. Data are shown as the mean of three measurements.

Protocol: Natalizumab ADA bridging ELISA protocol

Demonstration of Antibody Specificity

In figure 4, a microtiter plate was coated overnight with various antigens at a concentration of 5 μg/ml. After washing and blocking with PBST + 5% BSA, detection was performed using human Anti-Natalizumab Antibody, clone AbD21375 (HCA248) at a concentration of 2 μg/ml followed by HRP conjugated Rat Anti-DYKDDDDK Tag Antibody (MCA4764P, 1:500 dilution) in HISPEC Assay Diluent (BUF049A) and QuantaBlu Fluorogenic Peroxidase Substrate. 

Licensed Use: For in vitro research purposes and for commercial applications for the provision of in vitro testing services to support preclinical and clinical drug development. Any re-sale in any form or any other commercial application needs a written agreement with Bio-Rad.