SPR analysis was used to detect the binding between the recombinant ZSWIM4 protein (Cat. TSTG2502, TargetMol) and IPN60090, with the ZSWIM4 protein and anti-ZSWIM4 antibody combination used as the positive control. The anti-ZSWIM4 antibody and IPN60090 were diluted to different concentrations using the running buffer (5% DMSO). The ZSWIM4 protein was coupled to the CM5 sensor chip (GE Healthcare, Marlborough, MA, USA) and served as the ligand. SPR experiments were performed using a BIAcore T200 instrument (GE Healthcare) at 25 °C following the manufacturer’s instruction. The BIAcore T200 Control software (v. 2.0, GE Healthcare) was used to analyze the binding model.
Biacore t200 control software
Manufactured by GE Healthcare
Sourced in Australia
The BIAcore T200 Control software is a key component of the BIAcore T200 system, a widely used label-free interaction analysis instrument. The software serves as the primary interface for users to control the system's various functions, collect data, and analyze the results of biomolecular interaction studies.
Automatically generated - may contain errors
Lab products found in correlation
Cm5 sensor chip, by GE Healthcare (2 mentions)
Biacore t200, by GE Healthcare (2 mentions)
Methanol, by Thermo Fisher Scientific (2 mentions)
Dimethyl sulfoxide (dmso), by Thermo Fisher Scientific (2 mentions)
Biacore t200 instrument, by GE Healthcare (1 mentions)
Biacore t200 evaluation software version 1, by GE Healthcare (1 mentions)
Biacore t200 evaluation software, by GE Healthcare (1 mentions)
Prism v7, by GraphPad (1 mentions)
Sulfo nhs, by Bio-Rad (1 mentions)
3 protocols using biacore t200 control software
1
ZSWIM4 Protein-IPN60090 Binding Kinetics
2
Measuring G-CSF Variant Binding Kinetics
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The binding of G‐CSF variants to the G‐CSF receptor was measured by surface plasmon resonance (SPR) using a Biacore T200 (GE Healthcare). First, Protein A was immobilized on the CM5 sensor chip (GE Healthcare) via amine coupling in an immobilizing buffer containing 10 mm sodium acetate (pH 5.0). Purchased Fc‐chimera G‐CSF receptor was injected and captured via Protein A in running buffer (10 mm HEPES, 0.05% Tween20, 150 mm NaCl, pH 7.4). To measure the interaction with the receptor, each G‐CSF variant was sequentially injected into a flow cell in a single cycle manner at increasing concentrations and a flow rate of 30 μL·min−1. To measure the next variant, the sensor chip was regenerated by removing the G‐CSF receptor using a solution containing 10 mm glycine‐HCl (pH 2.0). Kinetic constants, including kon, koff, and KD, were calculated using software supplied with the instrument (Biacore T200 Control Software and Biacore T200 Evaluation Software, version 1.0, GE Healthcare).
3
Surface Plasmon Resonance Screening
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Commercially available reagents were used without additional purification unless otherwise stated. The fragment library (1000 wells, each containing 1 mg of a fragment compound) was purchased from Life Chemicals (Ontario, Canada). Phosphate-buffered saline (PBST; pH 7.4 with 0.05% Tween 20) was purchased from Sigma-Aldrich (Sydney, Australia). SPR reagents (40 mM EDC, 10 mM sulfo-NHS, 10 mM sodium acetate, 1 M ethanolamine HCl, pH 8.5) were purchased from Bio-Rad Laboratories (Gladesville, Australia). Methanol and DMSO were purchased from Thermo Fisher Scientific (Scoresby, Australia).
The SPR screening protocol was prepared with the Biacore T200 control software, measurements were performed using the Biacore T200 with a CM5 sensor chip, and the results were analyzed with the Biacore T200 evaluation software (GE Healthcare Life Sciences, Sydney, Australia).
The IC 50 , reported with standard error, was determined using GraphPad Prism v7.02 software (GraphPad, La Jolla, CA), by developing a nonlinear regression fit of the data collected from the inhibition assays. Images of docked compounds were generated by the PyMOL Molecular Graphics System (Schrodinger LLC, Cambridge, MA).
The SPR screening protocol was prepared with the Biacore T200 control software, measurements were performed using the Biacore T200 with a CM5 sensor chip, and the results were analyzed with the Biacore T200 evaluation software (GE Healthcare Life Sciences, Sydney, Australia).
The IC 50 , reported with standard error, was determined using GraphPad Prism v7.02 software (GraphPad, La Jolla, CA), by developing a nonlinear regression fit of the data collected from the inhibition assays. Images of docked compounds were generated by the PyMOL Molecular Graphics System (Schrodinger LLC, Cambridge, MA).
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