A whitepaper by Abveris
In vivo antibody discovery capitalizes on the natural immune repertoire of host animals to provide diverse, high affinity, and developable lead candidates suitable for downstream development.
However, the new wave of challenging therapeutic target classes are becoming too difficult for conventional hybridoma-based methods using wild-type mice.
To address the rising discovery challenges, Abveris has built a multi-faceted, accelerated discovery approach based on proprietary hyperimmune mice (DiversimAb™ family) and a Beacon-based single B cell screening platform that offers several benefits over traditional methods:
DiversimAb is a family of proprietary hyperimmune mouse models designed for expanded epitopic and sequence diversities (Figure 1).
DiversimAb mice break immune tolerance and show robust immune responses against many antigens including DNA, cell, and/or recombinant proteins, making them ideal workhorses for discovery targeting high-homology targets, cell surface receptors, and multi-pass transmembrane proteins such as GPCRs.
DiversimAb mice have been proven effective in the identification of species cross-reactivity, diverse affinities, and mouse-anti-mouse surrogates (Figure 2 & 3).
Figure 1. Illustration of the Enhanced Epitopic Coverage of mAbs Generated by DiversimAb Mice vs. Wild-Type Mice
Figure 2. Cross-Reactive Hits by Mouse Platform Against a Cell Surface Target X
Figure 3. 66 Discovery Campaigns Using DiversimAb Demonstrated Diverse Affinities Down to Single-Digit and Subnanmolar Ranges
The Beacon platform uses optofluidic technology to precisely manipulate the entrapment of single B cells into individual nanopens within a microfluidic chip. Assays are then performed on-chip using fluorescence imaging. The Beacon screening workflow is generally described in three stages (Figure 4):
(01) The process starts with loading of single B cells into a Beacon chip containing 14K individual nanopens. Screening reagents are then distributed throughput the channels where assays take place.
(02) Antibodies secreted by each B cell diffuse into the opening of a nanopen. Antibody binding to beads or cells is monitored by fluorescent readouts (“blooms”), allowing for the identification of individual hits. The Beacon platform can perform up to 4 multiplexed, sequential assays in just 1 day.
(03) Single B cells with desired activity profiles are then exported into a PCR plate for cDNA synthesis and single cell sequencing to recover the VH/VL paired sequences. Antibodies are then recombinantly expressed and validated for binding and activity off-Beacon.
Combining DiversimAb with the Beacon-based screening workflow enables:
Figure 4. Overview of a Beacon-Based Single B cell Workflow
Figure 5. The combination of DiversimAb mice in a Beacon-based workflow outperformed wild-type mice in a conventional hybridoma workflow in a discovery campaign against a cell surface receptor.
Two cohorts of DiversimAb mice were immunized following a rapid immunization schedule using recombinant protein of the FM1 extracellular domain as the immunogen and an Abveris proprietary adjuvant for maximum immune response.
On Day 18, sera were collected from the mice and titer tests were performed to examine target-specific immune responses to determine the best mice for tissue harvest on Day 21.
Viable plasma B cells were isolated from each mouse and imported into two separate Beacon chips using a proprietary loading protocol.
21,817 single B cells were screened to identify FM1-specific antibodies with the desired activity profile using three sequential assays (Figure 6). 276 hits of exhibiting on-chip ligand blocking & FM1-specific binding were identified.
Figure 6. Beacon assay cascade and representative results for the identification of FM1-specific, on-cell binders with ligand blocking activity
WHO WE ARE: As a division of Twist Biopharma, Abveris strives to provide exceptional in vivo antibody discovery services across the board, from project design and communication to final product delivery.
TRACK RECORD OF SUCCESS:
Abveris discovery platform advantages
