Anti-Ustekinumab Antibodies

Ideal for bioanalytical assays to measure the levels of ustekinumab and biosimilar products in preclinical research and clinical trials

Ready-made recombinant, anti-idiotypic monoclonal antibodies, highly specific for the human antibody drug ustekinumab (STELARA®)

Anti-Ustekinumab Type 1 Antibodies

Anti-Ustekinumab Type 1 Antibodies

The Type 1 anti-ustekinumab antibodies inhibit the binding of the drug ustekinumab to its target, IL-12 and IL-23, and therefore detect free drug. These antibodies are ideal for use in direct ELISA, pharmacokinetic (PK) bridging assays and to develop and calibrate immune response (IR) assays to measure the anti-drug antibody (ADA) response in patient sera.

The recombinant anti-ustekinumab antibodies are generated using the HuCAL® antibody library and a modified and improved form of phage display. Antibody generation using HuCAL technology results in highly specific and sensitive antibodies, which are ideal for PK and PD assays. Antibodies are fully human, so are also suitable as controls or calibrators for ADA assays. 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


Antibodies Specific to Ustekinumab

Product Code Clone Antibody Specificity Binding Type Format Affinity KD, nM Assay Development Recommendations
HCA208 AbD17829 Ustekinumab Type 1 Fab-V5Sx21 2.8 PK bridging ELISA capture antibody with HCA210
HCA209 AbD18334 Ustekinumab Type 1 Fab-FH2 0.4 PK bridging ELISA capture antibody with HCA210
HCA210 AbD17827_hIgG1 Ustekinumab Type 1 Human IgG1 0.2* PK bridging ELISA detection antibody
ADA assay
HCA210P AbD17827_hIgG1 Ustekinumab Type 1 Human IgG1
HRP labeled
0.2* PK bridging ELISA detection antibody
HCA211 AbD17829_hIgG1 Ustekinumab Type 1 Human IgG1 2.8* ADA assay

Table 1: Antibody Specifications
1 Fab antibody; V5-tag; StrepX-StrepX-tag
2 F=DYKDDDDK-tag; H=His-6-tag
*Affinity measured in the monovalent Fab format


Pharmacokinetic Assay - Bridging ELISA Format

Schematic image of anti-idiotypic antibodies in PK 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

PK bridging ELISA using antibodies HCA209 and HCA210

Figure 1: PK bridging ELISA using antibodies HCA209 and HCA210

Human anti-ustekinumab antibodies, product codes HCA209 and HCA210

Figure 1: Anti-ustekinumab antibody, clone AbD18334 (HCA209), was coated at 1.0 µg/ml on a microtiter plate and left over night. Washing and blocking was performed with 5% BSA in PBST.  Ustekinumab, titrated into 10% human serum, was then added.  Detection was performed by adding HRP conjugated anti-ustekinumab antibody, clone AbD17827_hIgG1 (HCA210P), 2 µg/ml in  HISPEC assay diluent (BUF049), plus QuantaBlu™ fluorogenic peroxidase substrate. The data are shown as the mean of three measurements.

Protocol: PK bridging ELISA for use with anti-ustekinumab antibodies HCA208 and HCA210P

Protocol: PK bridging ELISA for use with anti-ustekinumab antibodies HCA209 and HCA210P


Anti-Drug Antibody Assay - Bridging ELISA

Schematic image of ADA bridging assay.

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)

Immunogenicity assay, bridging format using antibodies HCA210 and HCA211

Figure 2: ADA assay, bridging format using antibodies HCA210 and HCA211

Human anti-ustekinumab antibodies, product codes HCA210 and HCA211

Figure 2: Ustekinumab was coated on a microtiter plate and left over night; coating concentration for the experiment using product HCA210 as reference standard was 1 µg/ml, and using product HCA211, 1.5 µg/ml. Washing and blocking was performed with 5% BSA in PBST. Anti-ustekinumab antibody, titrated into 10% human serum in the given concentrations, was then added. Detection was performed by adding HRP conjugated ustekinumab* (4 µg/ml in HISPEC asasy diluent), plus QuantaBlu fluorogenic peroxidase substrate. Data are shown as the mean of three measurements.

Protocol: ADA bridging ELISA for use with anti-ustekinumab antibody

*Antibody conjugated to HRP using a LYNX Rapid Conjugated Kit® (product codes LNK001P - LNK006P)


Inhibition ELISA

 figure 3 is the mean of three measurements. 

Figure 3: Inhibition of ustekinumab to IL-23 by antibody HCA209

 

Inhibition of ustekinumab binding to IL-23, product code HCA209

IL-23 (1.0 µg/ml) was used as the coating antigen.  A pre-incubated mixture of the anti-ustekinumab antibody, clone AbD18334 (HCA209) was titrated into a constant amount of ustekinumab (0.3 µg/ml).

Free ustekinumab still capable of binding to the plate was detected using HRP conjugated mouse anti-human IgG (Fc) CH2 domain (MCA647P).

The data are shown as the mean of three measurements. 


Determination of Antibody Specificity


Figure 4: Specificity of anti-ustekinumab antibody HCA211

Human anti-ustekinumab antibody, product code HCA211

Figure 4: Demonstration of the specificity of antibodies to ustekinumab. Antigens were coated at 5 µg/ml on a microtiter plate over night.  After washing and blocking with 5% milk powder in PBST, HRP conjugated anti-ustekinumab, clone AbD17829_hIgG1 (HCA211), 20µl from a 2µg/ml solution in HISPEC assay diluent, was added.  Detection was performed using 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.

STELARA® is a registered trademark of Janssen Biotech, Inc. USA. HuCAL® is a registered trademark of MorphoSys AG. QuantaBlu™ is a trademark of Thermo Fisher Scientific.