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Updated: May 23, 2026
A Fluorescence-based Lymphocyte Assay Suitable for High-throughput Screening of Small Molecules
Published on: March 10, 2017
Feasibility of implementing cell-based pathway reporter assays in early high-throughput screening assay cascades for
Alison J Smith1, Michael K Hancock, Kun Bi
1MedImmune Ltd, Cambridge, UK. SmithAJ@medimmune.com
Abstract:
Implementing functional cell-based screens in early antibody discovery has become increasingly important to select antibodies with the desired profile. However, this is limited by assay tolerance to crude antibody preparations and assay sensitivity. The current study aims to address this challenge and identify routes forward. Two common types of high-throughput screening (HTS) antibody sample, derived from either phage display or hybridoma techniques, have been screened across a wide range of CellSensor beta-lactamase reporter assays in a variety of cell backgrounds to more extensively characterize assay tolerance. Pathway-, sample-, and cell background-specific effects were observed. Reporter assays for agonism were less affected by crude antibody preparations, with 8 of 21 sample tolerant, and the potential to implement an additional 8 assays by choosing the best-tolerated sample type. Antagonist mode assays exhibited more complexity, with potentiating as well as inhibitory effects. However, 5 of 24 antagonist assays were fully tolerant, with the potential to implement an additional 11 assays. Different subsets of assays were affected in agonist versus antagonist mode, and hybridoma sample sets were better tolerated overall. The study clearly demonstrates the potential to use cell-based reporter assays in biologics HTS, particularly if the method of antibody production is considered in the context of the required assay mode (agonist/antagonist).
Insights
Early antibody discovery benefits from functional cell-based screening. This study characterizes assay tolerance to crude antibody preparations, finding hybridoma samples better tolerated and identifying routes for improved biologics high-throughput screening (HTS).
Area of Science:
- Biotechnology
- Immunology
- Drug Discovery
Background:
- Functional cell-based screening is crucial for antibody discovery.
- Assay tolerance to crude antibody preparations and sensitivity are key limitations.
- High-throughput screening (HTS) requires robust and adaptable assay systems.
Purpose of the Study:
- To characterize the tolerance of CellSensor beta-lactamase reporter assays to crude antibody preparations from phage display and hybridoma techniques.
- To identify factors influencing assay tolerance, including pathway, sample type, and cell background.
- To determine the feasibility of implementing cell-based reporter assays in biologics HTS.
Main Methods:
- Screening of phage display and hybridoma antibody samples across various CellSensor beta-lactamase reporter assays.
- Evaluation of assay tolerance in different cell backgrounds for both agonist and antagonist modes.
- Analysis of pathway-, sample-, and cell background-specific effects on assay performance.
Main Results:
- Agonist assays showed higher tolerance (8/21 fully tolerant) compared to antagonist assays (5/24 fully tolerant).
- Hybridoma samples demonstrated better overall tolerance than phage display samples.
- Specific assay modes and cell backgrounds influenced tolerance, with distinct effects observed for agonist versus antagonist assays.
Conclusions:
- Cell-based reporter assays can be effectively implemented in biologics HTS.
- Considering antibody production methods (hybridoma vs. phage display) and assay mode (agonist/antagonist) is critical for successful implementation.
- This characterization provides a foundation for optimizing cell-based screening strategies in antibody discovery.
