AGILE R100 Kinetic Characterization System

Information-rich kinetic binding data lets you understand measures critical to determining clinical efficacy, but previous systems have limitations in sensitivity, sample needs, and labeling requirements. Agile R100 is the first personal assay system that makes it easy to get accurate label-free analysis for small molecules and proteins at your benchCharacterize your candidates earlier in the process – and reduce costly mistakes further down the line.

Easily gain sensitive label-free data including:

  • Kinetic binding data and analysis
  • Affinity ranking
  • Dissociation rates
  • Concentration measurements
Label-free Agile R100 kinetic binding data assay

AGILE R100 FEATURES

  • Label-free platform simplifies your assay design, reduces confounding factors, and decreases costs.
  • Complex sample use enables detection in solvents such as 10% DMSO, detergents, cell fractions, tissue lysate, and serum, drastically reducing sample prep time.
  • 11-log dynamic range starting at 100 fM lets you develop weakly-binding fragments into high-affinity compounds on one platform.
  • Real-time data and single-sample format let you respond to immediate results to modify your assay mid-experiment.
  • 10 µL drop of sample so you can get more information from less material and reduce your cost to data.
  • Small molecule detection. Accurately detect molecules > 10 Da for flexible drug development unlimited by molecule size.
  • Measure unstable proteins. With no microfluidics, you can sense within minutes of sample prep, letting you measure labile interactions or unstable proteins quickly and easily.
  • Easy-to-learn. Because your sample is applied directly to the sensor surface, there’s no need to learn complicated components. If you can run an ELISA, you can run Agile R100.
  • Fits on your benchtop. Comprised solely of small, portable components, Agile R100 has no optical elements that cause other systems to be bulky and require calibration.
  • Breakthrough orthogonal technique that works when optical methods fail. See more about the Field Effect Biosensing technology on which Agile R100 is built.
How to use the Agile R100 system

APPLICATIONS

Kinetic binding data provides insight into your drug’s potential residence time, toxicity, and selectivity. Agile R100 lets you easily run assays at your bench in a free afternoon to characterize lead compounds and make confident decisions regarding your candidates.

Choose one of the below applications to learn more:

How to use the Agile R100 system

GETTING STARTED

This 7-minute overview video shows how to run a kinetic binding measurement on the Agile R100 system, the analysis that Agile Plus software provides, and the principles of Field Effect Biosensing (FEB) on which Agile R100 is based.

THE TECHNOLOGY DRIVING AGILE R100

At the heart of Agile R100 is a graphene biosensor chip built with proprietary label-free Field Effect Biosensing (FEB) technology. FEB is an electrical technique that measures the current across a biosensor surface. Only molecules binding to or dissociating from the biosensor surface cause a change in conductance, which the system reads and outputs as response. Unbound molecules or crude media that might interfere with optics do not affect the conductance reading. This robustness enables detection in complex samples from solvents and detergents to cell fractions and tissue lysate, making FEB a powerful technology for small molecule and protein drug discovery research.

Agile Biosensor Chip
Agile Plus software for Agile R100 label-free kinetic binding assay

AGILE PLUS SOFTWARE

Powerful, real-time Agile Plus software makes running assays and data analysis easy. Designed to work seamlessly with the single-sample Agile R100 platform, it is easy to learn and use. Preloaded protocols let you quickly set up your assay and run an experiment.The intuitive interface guides you through the process and automates the analysis so you can get to information-rich data faster. The real-time data display lets you visualize interactions as they occur.

  • Mid-experiment protocol modification lets you change your test in response to outputted data.
  • Automated analysis can be run at any time in the experiment for instant understanding of your results.
  • Pre-set PDF reporting options streamline data sharing.
  • Easy CSV data exporting supports additional analysis options on other software platforms.
  • No limit on software installation provides flexibility of use across computers.
Agile Plus software for Agile R100 label-free kinetic binding assay

THE AGILE R100 GETTING STARTED KIT

Tired of having to take a costly three-day course before you can start taking the data you need? You can learn everything you need to know about Agile R100 in four hours. The Agile R100 Getting Started Kit (GSK) is a self-training package that can be run in a free afternoon from the comfort of your bench, and it comes with every Agile R100 system. The GSK is a combination between 1) written guide and 2) experiment protocol and reagents that provides everything you need to run a TNFα protein and SPD304 small molecule interaction on the Agile R100 with confidence. In the GSK, you’ll find answers to questions such as, “What assay parameters can I change without messing up the data?”, “How do I know if my KD is good?” and “What is KD, anyway?” And it guides you through running a measurement on the Agile R100 and understanding the results. Think of it as your own personal three-day training package… that takes four hours.

Agile R100 label-free kinetic binding data

THE AGILE R100 GETTING STARTED KIT

Tired of having to take a costly three-day course before you can start taking the data you need? You can learn everything you need to know about Agile R100 in four hours. The Agile R100 Getting Started Kit (GSK) is a self-training package that can be run in a free afternoon from the comfort of your bench, and it comes with every Agile R100 system. The GSK is a combination between 1) written guide and 2) experiment protocol and reagents that provides everything you need to run a TNFα protein and SPD304 small molecule interaction on the Agile R100 with confidence. In the GSK, you’ll find answers to questions such as, “What assay parameters can I change without messing up the data?”, “How do I know if my KD is good?” and “What is KD, anyway?” And it guides you through running a measurement on the Agile R100 and understanding the results. Think of it as your own personal three-day training package… that takes four hours.

COMPONENTS

Graphene AGILE R100 biosensor chip

Biosensor Chips

Built with breakthrough Field Effect Biosensing (FEB) technology, Agile R100 biosensor chips help you fully characterize and explore your interactions to make better decisions earlier. The surface of the biosensor chip is made of graphene, platinum, and glass – biologically inert materials that do not react with your experiment and cause unnecessary nonspecific binding. Only molecules binding to or dissociating from the biosensor surface cause a change in conductance, enabling detection in complex samples from solvents and detergents to cell fractions and tissue lysate.

Agile biosensor chips are reusable and cost-effective, providing up to 10 measurements each. Unlike any other label-free platform, each measurement taken with an Agile biosensor chip provides three replicate data points for strong technical control.
See the Agile biosensor selection.

 

Label-free Agile R100 cartridge

Cartridge

The Agile R100 reader (not shown) houses the majority of the system’s electronic components. The Agile R100 cartridge holds the biosensor chip and biological materials. It is easy to clean, and an included clear cover reduces liquid evaporation.

The cartridge can connect directly to the reader for compact use and can be separated from the reader post-experiment for easy decontamination with standard protocols.

AGILE R100 cartridge with an optional cable accessory

The cartridge can also attach to the reader via a one-meter-long high density cable, which is an optional accessory. This enables additional use of the cartridge in a wide range of experimental settings (ex: with a temperature controlled incubator or in a biosafety cabinet) that may otherwise damage the reader.

Label-free kinetic binding assay Agile R100 reader

Reader

The Agile R100 reader houses the majority of the platform’s electronic components and powers the system via a USB connection to a computer or laptop. The reader can connect directly to the cartridge for compact use or be separated via a one-meter-long high density cable, which is an optional accessory. This enables a wide range of experimental settings (ex: with a temperature controlled incubator or in a biosafety cabinet) by providing the ability to use the cartridge in conditions that may otherwise damage the reader.

WORKFLOW AND SENSORGRAM

Agile R100, built with proprietary Field Effect Biosensing (FEB) technology, lets you monitor the interaction between two molecules in real-time. One is immobilized to the biosensor surface, and the other is a free-in-solution sample applied directly to the surface via a pipettor. The sensorgram below plots the response versus time, and is typical of how a real-time measurement is represented with the Agile R100 system and software. First, the calibration read in buffer is shown. Then, you can view the immobilization of the target on the biosensor surface. When your analyte is introduced, the association between target and analyte is seen in the sensorgram as the interaction takes place. Off-rates are measured as the interaction is reversed and dissociation occurs.

See the tabs in the below table for step-by-step details!

WORKFLOW

Agile R100, built with proprietary Field Effect Biosensing (FEB) technology, lets you monitor the interaction between two molecules in real-time. One is immobilized to the biosensor surface, and the other is a free-in-solution sample applied directly to the surface via a pipettor. The sensorgram below plots the response versus time, and is typical of how a real-time measurement is represented with the Agile R100 system and software. First, the calibration read in buffer is shown. Then, you can view the immobilization of the target on the biosensor surface. When your analyte is introduced, the association between target and analyte is seen in the sensorgram as the interaction takes place. Off-rates are measured as the interaction is reversed and dissociation occurs.

How the Agile R100 label-free kinetic binding assay works
Association and dissociation on the Agile R100 kinetic binding assay
AGILE R100 Field Effect Biosensing sensorgram

Calibrate in buffer

At the heart of Agile R100 is a revolutionary biosensor chip built with graphene, a unique nanomaterial. The graphene, platinum, and glass that make up the chip are inert materials that do not react with your biology.

A current is sent through the Agile biosensor chip and surrounding solution to monitor for changes in conductance. When performed with buffer, this is the initial baseline data in the sensorgram.

Immobilize target to biosensor chip surface

As the target is immobilized, it changes how the current applied to the solution affects the graphene. Changes in any exposed charge groups or to the configuration of the target alters the conductance, and the change in conductance is captured in the output data in real time.

 

 

Capture the binding interaction 

When analyte bind to the immobilized target, it alters the energy of the binding pair, which changes the conductance that is then read and outputted by the Agile R100 platform. Agile R100 is highly sensitive even to small changes in the conductance of the graphene biosensor surface.

 

 

Gain off-rates

As the analyte dissociates from the target, the decrease in response can be viewed in real-time, providing off-rates.

Because Agile R100 uses FEB, an electrical technique rather than an optical one, it is an excellent orthogonal option to double-check your hits, especially when optical methods fail.

 

View real-time data

Agile R100 measures changes in conductance in real-time, providing accurate and specific kinetics, affinity, and concentration data. Agile Plus software works seamlessly with Agile R100 and enables you to monitor results for immediate answers as they occur.

 

 

Learn more about the proprietary FEB technology behind Agile R100.

SPECIFICATIONS
Dynamic Range of Detection100 fM - 10 mM
Sample Volume10-50 uL
Functionalization Amount0.5-500 ng of material
Temperature Range-20C to 100 C
Lower Sense of Time2 minutes
Upper Sense of Time8+ hours
Size of Immobilized Target1 Da-200 kDa
Size of Analyte in Solution1 Da-200 kDa
Sample TypeSmall molecules in ≤ 10% DMSO, cell & tissue lysate, blood fractions
Target ChemistryEDC/sNHS or His-tag linkage
Speed of Data CollectionUp to 420 points/min
Shelf Stable3-6 months, stored with desiccant
Number of MeasurementsUp to 10 per Agile biosensor chip
Computer SpecificationsPC with USB port, Windows 7 or later
Weight2 pounds
Product Dimensions9 in. x 3 in.

DOWNLOADABLE LITERATURE

Comparison Studies

Agile R100 versus MicroCal iTC200 Kinetic Binding Data

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Agile R100 is a graphene biosensor with unprecedented functionality, providing label-free, real-time kinetic binding data. As a novel technology, Agile R100 is compared to MicroCal iTC200 to demonstrate its benefits compared to a standard kinetic binding analysis tool, isothermal titration calorimetry (ITC). In this study, kinetic binding data between the two instruments is compared by measuring Rho guanosine triphosphate hydrolase enzyme (GTPase) interacting with GTPase activating protein (GAP). The association and dissociation binding rate (ka and kd, respectively) and dissociation constants (KD) are reported. Agile R100 has a precise binding affinity measurement (KD = 1.35 ± 0.06 µM), comparable to MicroCal iTC200 (KD = 2.7 ± 0.3 µM), while also obtaining ka and kd values. Additionally, Agile R100 uses up to 30 million times less sample material and up to 30 times less sample volume and can perform 2.5 times more measurements in a given workday compared to MicroCal iTC200.

Application Notes

Reproducible Kinetic Characterization of Small Molecule Compounds in Serum with Agile R100

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  • In this application note, we show that Agile R100 provides reproducible kinetic binding data for small molecules with no assay adjustment when transitioning from buffer to serum. We examine 3 low molecular weight compounds interacting with their targets – a CR GPCR, a cytokine, and an aptamer in both buffer and serum. All kinetic constants between buffer and serum are within a factor of 1.9.
  • Agile R100 leverages breakthrough FEB technology to easily sense in complex matrices, providing earlier information about the behavior of a drug in serum without extensive assay development and optimization.

Reproducible Characterization of GPCR and Small Molecule Compound Interactions Using Agile R100

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GPCRs are primary drug targets for therapeutic intervention in multiple disease models, but they are notoriously difficult to characterize due to their inherent instability upon purification. Prior kinetic binding techniques have substantial roadblocks in studying GPCRs, but in this application note, we present Agile R100’s success in quantifying these difficult compounds. Features of Agile R100 that enable successful detection GPCR interactions include: the ability to sense in complex samples containing detergents and solvents, a non-microfluidic format that allows the sample to be applied directly to the surface of the biosensor chip, temperature versatility, and low target concentration and small sample volume requirements.

Small Molecule Compounds in DMSO Binding with GPCR and TNFα

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In this application note, we present Agile R100 kinetic binding data (kon, koff, and KD) from the interaction of 3 small molecule compounds dissolved in 10% DMSO binding with their respective targets – 2 GPCRs solubilized in additional detergents and a cytokine protein.

Detecting Interactions of Small Molecules in 10% DMSO Using Agile R100

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Small molecules are beneficial as drug therapeutics but often have low solubility in aqueous solutions. Solvents such as dimethyl sulfoxide (DMSO) are necessary additives for small molecule solubility, but these solvents can interfere with optical biosensing platforms. In this technical note, Agile R100 successfully detects the interaction between the cytokine tumor necrosis factor alpha (TNFα) and the small molecule inhibitor SPD304 in 0%, 1%, 3%, and 10% DMSO. Because Agile R100 uses an electrical technique rather than an optical one, DMSO in buffer is compatible with the instrument and does not require solvent correction. Additionally, high concentrations of SPD304 (100 μM) were detected on Agile R100. Importantly, Agile R100 detects the association binding rate (ka) and the dissociation binding rate (kd) and can calculate the dissociation constant (KD) of the TNFα and SPD304 interaction in buffer with DMSO. Agile R100’s KD result (14.5 ± 1.8 μM in phosphate buffer saline [PBS] with 10% DMSO) is similar to a previously reported KD value (7.3 ± 0.5 μM in citrate phosphate buffer with 10% DMSO).

Affinity Ranking of SPD304, Evans Blue, and Trypan Blue to TNFα Using Agile R100

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Affinity ranking is critical to optimizing the relationship between a target protein and a potential drug compound during the hit-to-lead optimization phase of drug discovery. In this application note, Agile R100 characterizes the activity of the target protein TNFα interacting with the small molecule drug compounds, SPD304, Evans Blue, and Trypan Blue in order to rank order by affinity.