Effects of serum matrix on the molecular interactions between drugs and target proteins as revealed by GMR biosensor technology
Investigations of molecular interactions between drugs and biomolecules are indispensable for understanding the mechanism of drug actions and their pharmacokinetics/pharmacodynamics. However, analyses of protein interactions are typically performed in buffered salt solutions which lack many components that are present in the original biological matrix, and thus has significant effects on the binding behaviors observed. This limitation of not being able to conduct binding kinetic studies in relevant biological matrices is a result of using an inadequate technology. The standard technology for such studies is based on surface plasmon resonance (SPR) measurements where nonspecific binding due to blood proteins distorts the kinetic data for samples with more than 1% serum. Therefore, detailed kinetic studies in relevant biological matrices are unavailable.
In a recent study published in J. Pharm. Biomed. Anal. 198, 114015 (2021), Saito, et al., used the Giant Magneto Resistance (GMR) platform from MagArray to investigate the binding characteristics of three molecular pairs in a serum-based matrix compared with a buffer-only matrix. The pairs were: quercetin and cAMP-dependent protein kinase A (PKA), Infliximab and tumor necrosis factor alpha (TNFα), and Bevacizumab and vascular endothelial growth factor (VEGF). They found that the presence of serum had different effects on the binding kinetics of the pairs compared to buffer-only measurements. Protein pairs were not as affected as were the small molecules. Saito et al., attributed the differences to the hydrophobic and electrostatic characteristics of the drug molecule, target protein, and serum proteins, which could only be evaluated with the GMR technology. They concluded that the real-time monitoring of molecular interactions in a more relevant biological matrix, enabled by the GMR platform, was a powerful tool to investigate such complicated, yet real-life, molecular interactions. Additionally, the method would allow the analysis of any kind of effects by a third molecule on the interaction between two other molecules, for example, an inhibitor drug’s interactions between two kinds of proteins. Such capabilities expand the depth of drug discovery and characterization.
To learn more about how the MagArray GMR platform can also help your drug discovery and characterization efforts, click here, or contact Kalidip Choudhury, Ph.D., at MagArray Lifesciences firstname.lastname@example.org
Giant Magnetic Resonance (GMR) Assay Principles
Our platform utilizes a sandwich assay in which the target antigen is sandwiched between two antigens, one bound to the magnetic sensor and the other to a superparamagnetic nano particle. Under an external magnetic field, the nanoparticles magnetize and their presence or absence can be detected by the sensor. Using chips with 64 GMR sensors we show rapid multiplex protein detection with a liner dynamic range of over 5-6 orders of magnitude for a diverse range of biological fluids in real time.
*Based on standard LBA process with magnetic nanoparticles used as labels
Instrument and chip
GMR Multiplex Chip
- 80 individual sensors on each chip
- 8 chips with 640 sensors total
- Each sensor can be individually spotted
- 80 biomarkers with controls for the multiplex chip
GMR Reader Unit
- Rugged and reliable
- No optics meaning no calibration of lasers, mirrors etc
- No fluidics means no pumps, valves, etc
- Fully automated
- Hands free incubation
- Bar code system to minimize errors
- No wash necessary
- Flexible assay protocols
- Run time of 1 hour for most sensitive assay
- Run time of 5-10 minutes for express assays
MagArray has 5-6 orders of magnitude greater dynamic range and a 1000 times more sensitive than ELISA.
Compared to ELISA, the MagArray platform is capable of detecting CEA at significantly lower concentrations and at 5-6 orders of magnitude greater dynamic range utilizing the same Ab pair
Matrix insensitive detection in different buffers.
A panel of eight human tumor markers with BSA negative control and epoxy control indicates matrix insensitive detection when shifting from PBS to mouse serum to lysis buffer
Ligand Binding Assay
Our GMR platform is more sensitive than SPR and insensitive to buffer conditions
MagArray platform at least 1000 times more sensitive than Biacore/SPR
MagArray platform insensitive to pH and salinity*
The effect of matrix on protein binding between SPR and MagArray
Buffer: 0.01M HEPES, pH 7.4, 0.15M NaCl, 3mM EDTA
Regeneration: 10mM Gly-HCL, pH 2.5
Analyte: TSH, Ligand: Anti-TSH ab
Biacore X 100
- The same binding pair and buffer conditions were used to compare between MagArray and Biacore
- Plasma was used as a matrix and diluted by PBS to 80%, 50%, 10%, 1%, 0.1%
- MagArray platform kinetic measurements were not affected by matrix
- Biacore did not provide reliable results above 0.1% matrix
- Truly a transformational platform that helps to bridge the gap between in-vitro and in-vivo.
- Matrix insensitive technology for measuring biomolecules in blood, plasma, serum, saliva and urine.
- 1000 times more sensitive than leading SPR technologies in measuring protein-protein interactions.
- Partnership with Hitachi Hi-Technologies allows expansion of platform for unmet needs in biological assays.
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