Proteins are useful biomarkers for a wide range of applications such as cancer detection, discovery of vaccines, and determining exposure to viruses and pathogens. Here, we present a low-noise front-end analog circuit interface towards development of a portable readout system for the label-free sensing of proteins using Nanowell array impedance sensing with a form factor of approximately 35cm2. The electronic interface consists of a low-noise lock-in amplifier enabling reliable detection of changes in impedance as low as 0.1% and thus detection of proteins down to the picoMolar level. The sensitivity of our system is comparable to that of a commercial bench-top impedance spectroscope when using the same sensors. The aim of this work is to demonstrate the potential of using impedance sensing as a portable, low-cost, and reliable method of detecting proteins, thus inching us closer to a Point-of-Care (POC) personalized health monitoring system. We have demonstrated the utility of our system to detect antibodies at various concentrations and protein (45 pM IL-6) in PBS, however, our system has the capability to be used for assaying various biomarkers including proteins, cytokines, virus molecules and antibodies in a portable setting.
Label-free technologies for the detection of proteins have made significant advancements in the last decade and have shown tremendous potential to be used as Point-of-Care devices in the literature14,15,16. Electrochemical biosensing technologies have the benefit of inherently having relatively high sensitivity and the ability to be used with miniaturized hardware thus having a smaller form factor than other technologies such as mass spectrometry or optical biosensing17. Electrochemical biosensors can be broadly classified into potentiometric, amperometric, and impedance biosensors based on the electrical parameter being measured. Potentiometric biosensors employ ion-sensitive field effect transistors (ISFETS) and ion selective electrodes for measuring a change in the electric potential due to accumulation of ions as a result of an enzymatic reaction18. Wang et al. have demonstrated the use of potentiometric sensors for the detection of carcinoembryonic antigen (CEA) with a sensitivity of 2.5 ng/ml19. Amperometric biosensors measure minuscule changes in the electrical current that take place due to redox reactions. The working electrode in such sensors is usually made of an inert material covered with a biorecognition element20.
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A fully integrated wearable impedance cytometer with an online smartphone readout has been demonstrated a little while back26. Recently, a handheld portable platform using disposable nanopore strips has also been presented for the detection of HIV antibody levels in saliva27. In another paper28, the authors presented a dielectrophoretic-impedance based method for the detection and analysis of proteins, specifically Bovine Serum Albumin (BSA). Authors in another article29 showcase an electrochemical impedance-based sensor employing graphene-carbon nanotubes composite deposited on a glassy carbon electrode to detect the protein antigen. A universal antibody-modified nanocrystalline boron-doped diamond biosensor for direct detection of protein and viral particles at very low concentrations was discussed30. Researchers described a new paper based electrochemical impedance biosensor for label-free detection and quantification of human interferon-gamma (IFN-γ), a biomarker which plays an important role in tuberculosis susceptibility31. Another microfluidic device, utilizing bead-based capture chamber technology, for the quantification of IL-6 in human serum samples for sepsis stratification was presented32. Furthermore, in other studies33,34, researchers present electrochemical biosensors for the detection of IgG anti-Trypanosoma cruzi antibodies and IgG antibodies to Helicobacter pylori in human serum samples respectively. Table 1 presents a comparison of systems that detect proteins electrically. Although significant leaps in the prototyping and development of microfluidic electrochemical biosensors have been made in the laboratory and academic environment within the last decade, the translation of most of these devices to a Point-of-Care setting has been limited by practical constraints and challenges such as portability, cost, integration, and regulatory affairs35. Additionally, the use of electrochemical biosensors for complex physiological samples such as blood, serum, and saliva is challenging since high salt concentrations in these samples reduce the double layer thickness and result in charge screening. Therefore, an electrical system needs to be highly sensitive, portable, and must have the ability to work with biological specimens with high salt concentrations for potential use in a POC setting.
We present the design of a portable, low noise electronic readout system to be used in conjunction with nanowell array impedance sensors for label-free sensing of proteins. The designed system has a small footprint of 35cm2. We carried out a comprehensive noise analysis of the system by calculating and simulating the output voltage referred noise and comparing it with the actual noise exhibited by the system. We have demonstrated the ability of the system to quantify antibodies by using varying antibody concentrations. We also determined the utility of the system to detect proteins by repeated detection of 45 pM IL-6 three times. Although we used a relatively slow protocol for our experiments to ensure robustness, our system has the ability to perform protein detection with a sample-to-answer turnaround time of 10 min when used in a practical setting. Although this work highlights the application of this system to be used for protein detection, it can be potentially used to study and measure the binding kinetics of the antibodies and proteins. To achieve this using the current system, we need only make slight modifications to the experimental protocol and analysis of the data. We have used a laptop computer and a separate board for the ATMega328 (Linduino) for storing the data. This can be replaced with an onboard microprocessor and the result can be transmitted wirelessly to a smartphone via Bluetooth in a future version of the system as we have demonstrated previously for a portable, microfluidic impedance cytometer41. A 3D printed package for the device with compartments for the circuit, the microcontroller, and the sensor is also a potential area where improvement can be made for using the system at Point-of-Care.
Measure and load impedance response(s).Check Ignore measured phase if phase response is not reliable.Measure Re if needed and Dd if Sd is not known.Enter parameters and added mass [g] or sealed box volume [liters] to text boxes.Check Mms or BL if value is trusted and second impedance response measurement is not available.Press Calculate.Results are visible in Calculated parameters and Z model groups.Basic Z model parameters Z1k and Z10k, and Extended Z model parameters Le, Leb,Ke and Rss are detected from free air impedance response.Also Re is calculated with Dual added mass method.Leach Z model parameters K and n are detected from free air impedance response,and simulated Z response is visible in chart if K is checked.See tooltip of curves for more information.
Traced sound pressure points can be copied to clipboard with Edit -> Copy raw SPL.Traced impedance points can be copied to clipboard with Edit -> Copy raw Z.Phase angle is zero and decimal symbol defined in Control panel.
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