Supplementary Fig. 4: Hemagglutinin-binding kinetics of 441D6 Fab by biolayer interferometry.
From: Mosaic nanoparticle display of diverse influenza virus hemagglutinins elicits broad B cell responses
Binding kinetics measurements were carried out using biolayer interferometry with Octet HTX instrument (fortéBio). Recombinant hemagglutinin trimers were immobilized either on HIS1K or SA sensors through hexa-histidine tag or conjugated biotin, respectively. Fab 441D6 was prepared by introducing HRV3C cleavage site in the heavy chain hinge and digesting with HRV3C enzyme followed by Protein A affinity purification to remove Fc and uncleaved antibodies as described elsewhere7. Dilution series of Fab 441D6 was made in assay buffer (PBS with 1% BSA). All assays were performed in the Octet HTX instrument (fortéBio) at 30ºC with agitation at 1,000 rpm. Represented data contain measured sensograms (red) and global fitting with 1:1 binding model (black) generated by Octet Analysis software (version 9.0, fortéBio). Experiments were independently performed two times with similar results.
