A global benchmark study using affinity-based biosensors

Abstract

To explore the variability in biosensor studies, 150 participants from 20 countries were given the same protein samples and asked to determine kinetic rate constants for the interaction. We chose a protein system that was amenable to analysis using different biosensor platforms as well as by users of different expertise levels. The two proteins (a 50-kDa Fab and a 60-kDa glutathione S-transferase [GST] antigen) form a relatively high-affinity complex, so participants needed to optimize several experimental parameters, including ligand immobilization and regeneration conditions as well as analyte concentrations and injection/dissociation times. Although most participants collected binding responses that could be fit to yield kinetic parameters, the quality of a few data sets could have been improved by optimizing the assay design. Once these outliers were removed, the average reported affinity across the remaining panel of participants was 620 pM with a standard deviation of 980 pM. These results demonstrate that when this biosensor assay was designed and executed appropriately, the reported rate constants were consistent, and independent of which protein was immobilized and which biosensor was used.

Rapid screening of IgG quality attributes – effects on Fc receptor binding

“…32, and Rich et al . 33), and therefore, variability was found acceptable for the intended purpose of the method.…”

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

“…The OctetRED instrument, which uses BLI, is a high-throughput, label-free platform that allowed us to screen 2000 supernatants kinetically within 15 h. Unlike traditional SPR instruments, the OctetRED uses a plate-based dip-and-read configuration, does not require the use of microfluidics, and therefore is less sensitive to sample and buffer composition compared to traditional flow-based SPR instruments. 7,8 Screening results showed that all antibodies selected by ELISA were also selected using the OctetRED platform. However, we identified several antibodies using the OctetRED that were missed in ELISA.…”

“…7 Traditional label-free platforms, such as the Biacore (GE Healthcare, Little Chalfont, UK), use surface plasmon resonance (SPR) coupled with a sophisticated microfluidics to flow analyte over ligand-coated sensors. 8 These traditional flow-based and SPR-based platforms have been previously used to kinetically screen hybridoma supernatants; however, the throughput is not very high (fewer than 800 samples a day). 9,10 The OctetRED uses a non-flow-based dip-and-read platform that can measure 16 samples in parallel by immersing ligand-coated biosensor tips into sample wells of a 96-or 384-well microplate.…”

Complementary Use of Label-Free Real-Time Biosensors in Drug Discovery of Monoclonal Antibodies

“…Determining whether there is consensus across different biosensor platforms operated by independent users has been a theme of several benchmark studies (Rich et al, 2009;Navratilova et al, 2007;Katsamba et al, 2006;Papalia et al, 2006;Cannon et al, 2004;Myszka et al, 2003). The largest and most recent of these engaged 150 participants from 20 countries who used instruments from ten different manufacturers.…”