Fortebio octet k2

A PCR amplifier was bought from Thermo Fisher (Waltham, MA, USA). Electrophoresis was implemented by a Bio-Rad apparatus (Hercules, CA, USA). An automatic microplate reader (SPARK) was bought from TECAN (Männedorf, Switzerland). An Odyssey^® Imaging system was from LI-COR (Lincoln, NE, USA). A molecular interaction analyzer (ForteBio Octet K2) was bought from SARTORIUS (Gottingen, Germany). ATR–FTIR was bought from Thermo (Waltham, MA, USA).

Binding kinetics data were obtained using a ForteBio Octet K2 instrument (Sartorius, Göttingen, Germany). The assays were performed at 30 °C in 96-well black plates. Biotinylation of daclizumab (#MAB9927, R&D Systems) was performed using a biotin labeling kit (Dojindo, Kumamoto, Japan) in accordance with the manufacturer’s instructions. Biotinylated daclizumab or Zif-FLAP were loaded onto the surface of a streptavidin biosensor (Sartorius) at 100 nM in kinetic buffer (0.1% BSA and 0.002% Tween-20 in TBS, pH 7.5) for 300 s. After washing (30 s) and equilibrating (60 s) the biosensor, the association of the ligand on the biosensor to CD25-Fc (ACROBiosystems, Newark, DE, USA) was measured for 300 s. The biosensor was then dipped back into buffer for another 300 s to measure dissociation. Systematic baseline drift correction was conducted by subtracting the shift recorded for sensors loaded with ligand, but incubated without analyte. Data analysis and curve fitting were performed using Octet software version 11.0. Experimental data were fitted with the binding equations available for a 1:1 interaction with local fitting to calculate K_D, k_on, and k_off values using data obtained from four analyte (CD25-Fc) concentrations.

Phagemid vector pHB was from the Academy of Military Medical Sciences. SfiI, NotI enzymes, and M13KO7 helper phages were bought from New England Biolabs (Ipswich, MA, USA). ELISA coating buffer and TMB substrate solution were bought from Solarbio. Rabbit anti-SARS-CoV-2 S protein polyclonal antibody was bought from Sino Biological (No. 40592-T62-100) (Beijing, China). Rabbit anti-M13 pAb-HRP was bought from Antaizhiyuan Technology Co., Ltd. (Beijing, China). Syringe Filter Units (0.45 µm) were bought from Merck Millipore (Burlington, MA, USA). Broths and culture media were prepared by ourselves. The automatic microplate reader (SPARK) was bought from TECAN. The molecular interaction analyzer (ForteBio Octet K2, Fremont, CA, USA) was bought from SARTORIUS. FLS1000 Fluorescence Spectrophotometer was from Edinburgh Instruments Ltd (Livingston, UK). Moreover, DXR3 Raman spectrometer was from ThermoFisher Inc (Waltham, MA, USA).