High-Throughput Kinetic Screening of Hybridomas to Identify High-Affinity Antibodies Using Bio-Layer Interferometry

Lad, Latesh; Clancy, Sheila; Коваленко, М. А.; Liu, Chian; Smith, Victoria; Pagratis, Nikos

doi:10.1177/1087057114560123

Cited by 39 publications

(29 citation statements)

References 31 publications

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“…This approach is costly, time-consuming, and restricts the number of clones that can be evaluated. Several work on the developments of high-throughput assessment of binding affinities has been reported and remained being assessed3738.…”

Section: Discussionmentioning

confidence: 99%

Continuous microfluidic assortment of interactive ligands (CMAIL)

Hsiao

Huang

et al. 2016

Sci Rep

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Finding an interactive ligand-receptor pair is crucial to many applications, including the development of monoclonal antibodies. Biopanning, a commonly used technique for affinity screening, involves a series of washing steps and is lengthy and tedious. Here we present an approach termed continuous microfluidic assortment of interactive ligands, or CMAIL, for the screening and sorting of antigen-binding single-chain variable antibody fragments (scFv) displayed on bacteriophages (phages). Phages carrying native negative charges on their coat proteins were electrophoresed through a hydrogel matrix functionalized with target antigens under two alternating orthogonal electric fields. During the weak horizontal electric field phase, phages were differentially swept laterally depending on their affinity for the antigen, and all phages were electrophoresed down to be collected during the strong vertical electric field phase. Phages of different affinity were spatially separated, allowing the continuous operation. More than 105 CFU (colony forming unit) antigen-interacting phages were isolated with ~100% specificity from a phage library containing 3 × 109 individual members within 40 minutes of sorting using CMAIL. CMAIL is rapid, sensitive, specific, and does not employ washing, elution or magnetic beads. In conclusion, we have developed an efficient and cost-effective method for isolating and sorting affinity reagents involving phage display.

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Section: Discussionmentioning

confidence: 99%

Continuous microfluidic assortment of interactive ligands (CMAIL)

Hsiao

Huang

et al. 2016

Sci Rep

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“…Biolayer interferometry was performed on a BlItz® System (Pall ForteBio LLC, CA) according to the description by Lad et al 32. Briefly, IgG1 Fc-fusion receptor proteins (PDGFRβ-Fc, DR4-Fc or DR5-Fc) were immobilized on a protein A-coated biosensor.…”

Section: Methodsmentioning

confidence: 99%

Targeted Delivery to Tumor-associated Pericytes via an Affibody with High Affinity for PDGFRβ Enhances the in vivo Antitumor Effects of Human TRAIL

Tao¹,

Yang²,

Shi³

et al. 2017

Theranostics

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Human tumor necrosis factor-related apoptosis-inducing ligand (hTRAIL) has exhibited superior in vitro cytotoxicity in a variety of tumor cells. However, hTRAIL showed a disappointing anticancer effect in clinical trials, although hTRAIL-based regimens were well tolerated. One important reason might be that hTRAIL was largely trapped by its decoy receptors, which are ubiquitously expressed on normal cells. Tumor-targeted delivery might improve the tumor uptake and thus enhance the antitumor effect of hTRAIL. Platelet-derived growth factor receptor β (PDGFRβ)-expressing pericytes are enriched in tumor tissues derived both from patients with colon cancer and from mice bearing colorectal tumor xenografts. A ZPDGFRβ affibody showed high affinity (nM) for PDGFRβ and was predominantly distributed on tumor-associated PDGFRβ-positive pericytes. Co-administration with the ZPDGFRβ affibody did not significantly enhance the antitumor effect of hTRAIL in mice bearing tumor xenografts. Fusion to the ZPDGFRβ affibody endows hTRAIL with PDGFRβ-binding ability but does not interfere with its death receptor binding and activation. The fused ZPDGFRβ affibody mediated PDGFRβ-dependent binding of hTRAIL to pericytes. In addition, hTRAIL bound on pericytes could kill tumor cells through juxtatropic activity or exhibit cytotoxicity in tumor cells after being released from pericytes. Intravenously injected hTRAIL fused to ZPDGFRβ affibody initially accumulated on tumor-associated pericytes and then diffused to the tumor parenchyma over time. Fusion to the ZPDGFRβ affibody increased the tumor uptake of hTRAIL, thus enhancing the antitumor effect of hTRAIL in mice bearing tumor xenografts. These results demonstrate that pericyte-targeted delivery mediated by a ZPDGFRβ affibody is an alternative strategy for tumor-targeted delivery of anticancer agents.

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“…Typically, a target will be used to immunize mice or used as a selection agent and the antibodies produced will be screened for desired properties. Although screens can be performed using antibodies in unpurified supernatants, as is often done with hybridoma campaigns [7], some assays are very sensitive to contaminants [8] and a form of purification is required. With the hundreds or even thousands of potential drug candidates being generated by display technologies, traditional methods impede the momentum of screening such a large number of hits due to the very nature of the purification protocols.…”

Section: Glycosylationmentioning

confidence: 99%

Current advances in the development of high-throughput purification strategies for the generation of therapeutic antibodies

Spooner¹,

Wilkinson²,

Kemp³

2015

Pharmaceutical Bioprocessing

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The demand for therapeutic proteins, and particularly monoclonal antibodies, continues to grow within the biopharmaceutical industry. This is illustrated by the monoclonal antibody Humira ® (adalimumab) having the highest sales of any drug in 2014. With the increasing numbers of biologics entering the market and the emergence of biosimilars and biobetters, costs need to be reduced in order for companies to remain competitive. High-throughput purification can provide the tools to meet the challenge of producing the thousands of potential drug candidates generated in drug discovery while simultaneously providing robust and cost-effective purification strategies in both research and production. In this review, we discuss the many different methodologies and automation technologies that can be employed in high-throughput purification.

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High-Throughput Kinetic Screening of Hybridomas to Identify High-Affinity Antibodies Using Bio-Layer Interferometry

Cited by 39 publications

References 31 publications

Continuous microfluidic assortment of interactive ligands (CMAIL)

Continuous microfluidic assortment of interactive ligands (CMAIL)

Targeted Delivery to Tumor-associated Pericytes via an Affibody with High Affinity for PDGFRβ Enhances the in vivo Antitumor Effects of Human TRAIL

Current advances in the development of high-throughput purification strategies for the generation of therapeutic antibodies

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