Off-Rate Ranking & Epitope Binning for Monoclonal Antibody Characterization

Advanced Bio-Layer Interferometry (BLI) Workflows for Antibody Discovery

The Challenge: Limited Screening Increases Development Risk

Traditional antibody development workflows often evaluate a relatively small number of clones and do not fully differentiate binding kinetics or epitope diversity across large candidate panels. When affinity and binding relationships are critical, this limited visibility can increase development risk and delay identification of fit-for-purpose reagents.

In many applications, dissociation rate (k_off) plays a key role in performance. For example, diagnostic assays such as ELISA rely on stable antibody–antigen interactions through multiple wash steps, while therapeutic antibodies often require slow dissociation to maintain target engagement. Without early insight into these properties, downstream performance can be difficult to predict.

The Solution: High-Throughput Off-Rate Ranking + Epitope Binning

Rockland has integrated high-throughput off-rate ranking and in-tandem epitope binning into its antibody discovery workflow using the Sartorius Octet® Bio-Layer Interferometry (BLI) platform. This approach enables rapid screening of dozens to hundreds of hybridoma clones, providing early insight into both binding kinetics and epitope diversity.

By combining these complementary techniques, large antibody panels can be efficiently triaged and characterized prior to full kinetic analysis, accelerating decision-making and reducing resource requirements.

Off-Rate Ranking: Rapid Antibody Triage Without Purification

Off-rate ranking by BLI enables rapid comparison of the relative binding behavior of antibody candidates against a common antigen. Because the k_off is independent of analyte concentration during the dissociation phase, reliable relative ranking can be achieved without precise antibody quantitation.

Importantly, off-rate measurements can be performed directly in crude hybridoma supernatants obtained from conditioned tissue culture media. This eliminates the need for purification or full kinetic characterization during early screening and allows large antibody cohorts (e.g., >100 clones) to be evaluated rapidly while minimizing sample consumption.

Sample Preparation Time

Eliminates purification requirements during early screening

Reagent Consumption

Minimal sample volume required for large-scale evaluation

Overall Screening Effort

Rapid triage of large antibody cohorts

Representative BLI sensorgram illustrating off-rate ranking across antibody candidates, where dissociation rates are used to compare relative binding strength.

Epitope Binning: Identifying Compatible Antibody Pairs

While off-rate ranking identifies strong binders, it does not determine whether antibodies recognize the same or different regions of an antigen. For many downstream applications—particularly sandwich ELISA, lateral flow assays, and other dual-antibody formats—epitope diversity is essential.

Rockland’s in-tandem epitope binning workflow uses sequential binding assays to evaluate competitive and non-competitive interactions between antibodies. This enables classification of antibodies into distinct epitope bins based on shared binding behavior, supporting identification of compatible antibody pairs and mapping of binding diversity across large panels.

Apply These Workflows to Your Antibody Programs

Rockland’s integrated antibody discovery and characterization capabilities extend beyond this study, enabling rapid, high-confidence evaluation of antibodies against a wide range of targets. These workflows can be applied at any stage of development to support screening, selection, and assay design.