The development of Neutralizing Antibodies (NAbs) detection can provide effective immunological information for biopharmaceuticals or related disease intervention strategies, and the specific assay format selection depends on a combination of three key factors: drug type and mechanism of action (MoA), immunogenicity risk, and assay performance in obtaining reliable data.
In addition to providing comprehensive and reproducible neutralizing antibody evaluation services to scientists worldwide, Creative Biolabs also aims to provide guidance and support for your bio-detection projects.
Table 1. Cell-based assay (CBA) and competitive ligand-binding assay (CLBA). (Creative Biolabs)
| MoA | Type | Target | Mechanism | Recommended Method |
| Antagonist | Monoclonal antibodies | Humoral receptors | The drug binds and activates the receptor | CLBA |
| Cell receptor | The drug binds to cellular receptors and competitively inhibits receptor binding to ligand | CBA or CLBA | ||
| Soluble receptor | Ligand | Soluble receptor binds ligand, blocking receptor-ligand interaction | CLBA is recommended, CBA is an option when suitable cell lines are available | |
| Antibody-mediated delivery of cytotoxins into cells | ADC | Cell receptor | Adcs bind cellular receptors and mediate endocytosis | CBA is recommended, but CLBA is acceptable with drug interference |
| Receptor agonist | Recombinant protein or antibody | Cell receptor | The drug binds and activates the receptor | Cba preferred |
| Enzyme replacement | Enzyme | Replaces proteins lacking in circulating or target cells | Enzymes function through cellular uptake | Enzyme activity assay and/or CBA |
| Lysis of target cells by antibody effects | Monoclonal antibodies | Target cell receptor, FcyR, or complement | Antibodies bind to target cell receptors through variable regions and FcyR or to complement through Fc domains | CBA is recommended, CLBA is also acceptable |
Cell-based analyses provide cell line-specific functional measurements, which can more accurately describe the overall physiological effects of NAbs in biological systems rather than limited information on NAbs-drug interactions. Cell-based NAbs functional analysis is FDA-recommended and is the preferred choice for certain studies with complex mechanisms of action.
Endpoints in cell-based NAbs analysis usually include cell proliferation, apoptosis, viability, enzyme activity, protein secretion, receptor phosphorylation, mitochondrial function, or cytotoxicity. Based on whether the drug directly acts on cells or stimulates a response, cell-based analyses can be further classified as direct or indirect detection.
Fig 1. Direct and indirect NAb assays. (Creative Biolabs)
If the MoA of the drug or biopharmaceutical is blocking soluble targets/cell receptors or if developing reliable cell-based assays is challenging, non-cell-based detection methods are considered appropriate. Non-cell-based analyses are relatively easier to design and establish, compatible with more detection systems (ELISA, ECL, etc.), and with higher analytical precision, detection sensitivity, and dynamic range. Depending on the selected signaling molecules for the detection endpoint, non-cell-based NAbs assays can also be further classified (Ligand Binding NAb Assays, Enzymatic Activity NAb Assays, etc.).
Table 2. Differences between cell-based and non-cell-based NAbs assays.
| Cell-based NAbs Functional Assays | Non-cell-based NAbs Assays | |
| Advantages |
Functional/biological activity test Closer to physiological conditions |
Stable with small variability Low development difficulty The experiment is simple and less time-consuming |
| Disadvantages |
Experimental variability Complex experiment design and operation Time-consuming |
Functional characterization not possible |
Our experienced team of developers and scientists always stays at the forefront of biotechnology and antibody research. Here, we provide further knowledge on NAbs that may interest you, aiming to support your research endeavors.
In addition to advanced research tools and antibody products, Creative Biolabs also provides comprehensive technical support. We will comprehensively evaluate the required assay format, cell line, NAbs, optimize assay parameters and processing/pre-processing procedures to offer customized NAbs assay development services with our cutting-edge recombinant antibody production platform, and provide optimized standardized NAbs functional evaluations.
For further information, collaboration inquiries, or any assistance, please feel free to contact us.
