Data were collected on a Biacore X100 (GE Healthcare). Experiments were performed at 25 °C in Dulbecco’s PBS + 0.005 % Polysorbate-20 (GE Healthcare). In Figure 1 and Supplementary Table 2 a sensor chip protein A (GE Healthcare) was used to capture 50-100 RU of purified mAb diluted in SPR running buffer at a flow rate of 5 μL/min on flow cell 2. Next, an appropriate range (typically 20 nM-0.625 nM) of six 2-fold dilutions, with one replicate, of PvDBPII (expressed in S2 cells as described above) was injected for 90 s at 60 μL/min and dissociation was measured for 1600 s (7200 s when necessary). Specific binding of the PvDBPII protein to mAb was obtained by reference-subtracting the response of a blank surface from that of the mAb-coated surface. The sensor surface was regenerated with a 60 s pulse of 10 mM glycine-HCl pH 1.5 (GE Healthcare). Sensorgrams were fitted to a global Langmuir 1:1 interaction model, allowing determination of the kinetic association and dissociation rate constants using Biacore X100 evaluation software.
Glycine hcl ph 1
Manufactured by GE Healthcare
Sourced in United States
Glycine-HCl pH 1.5 is a laboratory buffer solution with a pH of 1.5. It is commonly used as a standard solution for calibrating and verifying the performance of pH meters and other pH measurement equipment.
Automatically generated - may contain errors
Lab products found in correlation
Polysorbate 20, by GE Healthcare (2 mentions)
Biacore x100, by GE Healthcare (2 mentions)
Protein a sensor chip, by GE Healthcare (2 mentions)
Biacore t200 evaluation software version 3, by GE Healthcare (1 mentions)
Biacore t200, by GE Healthcare (1 mentions)
Protein g sensor chip, by GE Healthcare (1 mentions)
Hbs ep, by GE Healthcare (1 mentions)
Biacore t200, by Cytiva (1 mentions)
3 protocols using glycine hcl ph 1
1
Affinity Kinetics of Pv DBPII-mAb Interaction
2
Binding Kinetics of Monoclonal Antibodies to HSA
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The affinity and kinetic of Mab binding to HSA was determined by BiacoreTM T200 (GE Healthcare life sciences, Illinois, USA), a Surface Plasmon Resonance instrument. HBS‐EP+ (GE Healthcare life sciences, Illinois, USA) was used as a system running buffer. Goat anti‐mouse IgG1 (Sigma‐Aldrich, Missouri, USA) was immobilized on the surface of the Protein G sensor chip (GE Healthcare life sciences, Illinois, USA). All ten clones of MAbs (ligand) were diluted to the concentration of 3 µg/ml with running buffer. A concentration of 20 nM and 320 nM of HSA (analyte) was injected with a flow rate of 30 µl/min for 30 s. The Protein G sensor chip surface was regenerated with 10 mM Glycine HCl pH 1.5 (GE Healthcare life sciences, USA). The results were analyzed with BIAcore T200 evaluation software version 3.1 (GE Healthcare life sciences, USA).
3
Affinity Kinetics of Pv DBPII-mAb Interaction
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Data were collected on a Biacore X100 (GE Healthcare). Experiments were performed at 25 °C in Dulbecco’s PBS + 0.005 % Polysorbate-20 (GE Healthcare). In Figure 1 and Supplementary Table 2 a sensor chip protein A (GE Healthcare) was used to capture 50-100 RU of purified mAb diluted in SPR running buffer at a flow rate of 5 μL/min on flow cell 2. Next, an appropriate range (typically 20 nM-0.625 nM) of six 2-fold dilutions, with one replicate, of PvDBPII (expressed in S2 cells as described above) was injected for 90 s at 60 μL/min and dissociation was measured for 1600 s (7200 s when necessary). Specific binding of the PvDBPII protein to mAb was obtained by reference-subtracting the response of a blank surface from that of the mAb-coated surface. The sensor surface was regenerated with a 60 s pulse of 10 mM glycine-HCl pH 1.5 (GE Healthcare). Sensorgrams were fitted to a global Langmuir 1:1 interaction model, allowing determination of the kinetic association and dissociation rate constants using Biacore X100 evaluation software.
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