Abstract:Affinity‐based electrochemical (EC) sensors offer a potentially valuable approach for point‐of‐care (POC) diagnostics applications, and for the detection of diseases, such as sepsis, that require simultaneous detection of multiple biomarkers, but their development has been hampered due to biological fouling and EC noise. Here, an EC sensor platform that enables detection of multiple sepsis biomarkers simultaneously by incorporating a nanocomposite coating composed of crosslinked bovine serum albumin containing… Show more
“…Incorporating terminal amine groups in rGOx augments their interaction with polymer matrices and their distribution by altering their solubility and agglomeration, in addition to enhancing the physical, mechanical, thermal, and electrical stability of the composite via covalent linking of the amines by GA pyridine polymers. 16, 17 Indeed, when we compared the effects of amine-functionalized rGOx we used previously versus pentaamine-functionalized rGOx (prGOx), we found that the prGOx maintained a higher current density and lower peak-to-peak distance when measured immediately after coating or after incubating in 1% BSA for 1h or 1 day ( Fig. 1b ).…”
Section: Ultra-fast Deposition Of An Antifouling Coating On Ec Sensorsmentioning
confidence: 93%
“…To develop an efficient process flow for mass manufacturing, a nanocomposite coating method was developed using on-chip heating to facilitate rapid deposition of our previously described BSA/rGOx composite. 17 Clean gold electrodes were heated at 85 0 C with the nanocomposite material for 30 sec to 5 min, followed by washing in PBS immediately or after cooling down for 10 min. This ultra-rapid cross-linking mechanism results in creation of porous three dimensional (3D) molecular networks, and we were able to detect this reaction by measuring increase in absorbance at 265-270 nm (Supplementary Fig.…”
Section: Ultra-fast Deposition Of An Antifouling Coating On Ec Sensorsmentioning
confidence: 99%
“…We previously described a highly effective anti-fouling nanocomposite coating composed of denatured bovine serum albumin (BSA) cross-linked to gold nanomaterials or reduced graphene nanoparticles (rGOx) using glutaraldehyde (GA). 12, 17 While this method was simple and greatly suppressed signal noise due to biofouling, it took 24 hours to prepare these surfaces. Here, we report an alternative version of this nanocomposite coating that is composed of BSA cross-linked to pentaamine graphene flakes that can be coated on EC sensors using localized heating in the unprecedented time of less than one minute, which allows for outstanding signal transduction between the gold electrode and the nanocomposite surface.…”
Here we describe an ultra-fast (< 1 min) method for coating electrochemical (EC) sensors with an anti-fouling nanocomposite layer that can be stored at room temperature for months, which provides unprecedented sensitivity and selectivity for diagnostic applications. We leveraged this method to develop a multiplexed diagnostic platform for detection of biomarkers that could potentially be used to triage patients with myocardial infarction and traumatic brain injury using only 15 microliters of blood. Single-digit pg/mL sensitivity was obtained within minutes for all the biomarkers tested in unprocessed human plasma samples and whole blood, which is much faster and at least 50 times more sensitive than traditional ELISA methods, and the signal was stable enough to be measured after one week of storage. The multiplexed EC sensor platform was validated by analyzing 22 patient samples, which demonstrated excellent correlation with reported clinical values.
“…Incorporating terminal amine groups in rGOx augments their interaction with polymer matrices and their distribution by altering their solubility and agglomeration, in addition to enhancing the physical, mechanical, thermal, and electrical stability of the composite via covalent linking of the amines by GA pyridine polymers. 16, 17 Indeed, when we compared the effects of amine-functionalized rGOx we used previously versus pentaamine-functionalized rGOx (prGOx), we found that the prGOx maintained a higher current density and lower peak-to-peak distance when measured immediately after coating or after incubating in 1% BSA for 1h or 1 day ( Fig. 1b ).…”
Section: Ultra-fast Deposition Of An Antifouling Coating On Ec Sensorsmentioning
confidence: 93%
“…To develop an efficient process flow for mass manufacturing, a nanocomposite coating method was developed using on-chip heating to facilitate rapid deposition of our previously described BSA/rGOx composite. 17 Clean gold electrodes were heated at 85 0 C with the nanocomposite material for 30 sec to 5 min, followed by washing in PBS immediately or after cooling down for 10 min. This ultra-rapid cross-linking mechanism results in creation of porous three dimensional (3D) molecular networks, and we were able to detect this reaction by measuring increase in absorbance at 265-270 nm (Supplementary Fig.…”
Section: Ultra-fast Deposition Of An Antifouling Coating On Ec Sensorsmentioning
confidence: 99%
“…We previously described a highly effective anti-fouling nanocomposite coating composed of denatured bovine serum albumin (BSA) cross-linked to gold nanomaterials or reduced graphene nanoparticles (rGOx) using glutaraldehyde (GA). 12, 17 While this method was simple and greatly suppressed signal noise due to biofouling, it took 24 hours to prepare these surfaces. Here, we report an alternative version of this nanocomposite coating that is composed of BSA cross-linked to pentaamine graphene flakes that can be coated on EC sensors using localized heating in the unprecedented time of less than one minute, which allows for outstanding signal transduction between the gold electrode and the nanocomposite surface.…”
Here we describe an ultra-fast (< 1 min) method for coating electrochemical (EC) sensors with an anti-fouling nanocomposite layer that can be stored at room temperature for months, which provides unprecedented sensitivity and selectivity for diagnostic applications. We leveraged this method to develop a multiplexed diagnostic platform for detection of biomarkers that could potentially be used to triage patients with myocardial infarction and traumatic brain injury using only 15 microliters of blood. Single-digit pg/mL sensitivity was obtained within minutes for all the biomarkers tested in unprocessed human plasma samples and whole blood, which is much faster and at least 50 times more sensitive than traditional ELISA methods, and the signal was stable enough to be measured after one week of storage. The multiplexed EC sensor platform was validated by analyzing 22 patient samples, which demonstrated excellent correlation with reported clinical values.
“…Several recent reports on multiplexed sensors with PCT as one of the analytes are available in literature [48]. A lab-on-a-chip immunosensor array was reported for simultaneous detection of the pro-inflammatory markers PCT, interleukin-6 (IL-6) and C-reactive protein (CRP) using a cyclooctene (TCO)-tetrazine (Tz)-horse radish peroxidase (HRP) assembly [49]. The ratio of TCO-HRP and Tz-HRP was optimized to achieve adequate chemiluminescent response for the different markers that are present in varying levels in blood.…”
Human procalcitonin (PCT) is a peptide precursor of the calcium-regulating hormone calcitonin. Traditionally, PCT has been used as a biomarker for severe bacterial infections and sepsis. It has also been recently identified as a potential marker for COVID-19. Normally, serum PCT is intracellularly cleaved to calcitonin, which lowers the levels of PCT (<0.01 ng/mL). In severe infectious diseases and sepsis, serum PCT levels increase above 100 ng/mL in response to pro-inflammatory stimulation. Development of sensors for specific quantification of PCT has resulted in considerable improvement in the sensitivity, linear range and rapid response. Among the various sensing strategies, electrochemical platforms have been extensively investigated owing to their cost-effectiveness, ease of fabrication and portability. Sandwich-type electrochemical immunoassays based on the specific antigen–antibody interactions with an electrochemical transducer and use of nanointerfaces has augmented the electrochemical response of the sensors towards PCT. Identification of a superior combination of electrode material and nanointerface, and translation of the sensing platform into flexible and disposable substrates are under active investigation towards development of a point-of-care device for PCT detection. This review provides an overview of the existing detection strategies and limitations of PCT electrochemical immunosensors, and the emerging directions to address these lacunae.
“…Here we demonstrate the sensitivity, repeatability and specificity of the GFET biosensors for the detection of pure proteins. The next stage of the study will be to investigate the sensor response using real patient samples (such as serum, plasma, or blood), stability and storage of the sensors (Teixeira et al, 2014;Leva-Bueno et al, 2020;Zupančič et al, 2021). Thus, GFET biosensors offer a unique route towards the development of vitally needed diagnostic platform for AD.…”
We report on the fabrication and characterisation of graphene field-effect transistor (GFET) biosensors for the detection of Clusterin, a prominent protein biomarker of Alzheimer’s disease (AD). The GFET sensors were fabricated on Si/SiO2 substrate using photolithographic patterning and metal lift-off techniques with evaporated chromium and sputtered gold contacts. Raman Spectroscopy was performed on the devices to determine the quality of the graphene. The GFETs were annealed to improve their performance before the channels were functionalized by immobilising the graphene surface with linker molecules and anti-Clusterin antibodies. Concentration of linker molecules was also independently verified by absorption spectroscopy using the highly collimated micro-beam light of Diamond B23 beamline. The detection was achieved through the binding reaction between the antibody and varying concentrations of Clusterin antigen from 1 to 100 pg/mL, as well as specificity tests using human chorionic gonadotropin (hCG), a glycoprotein risk biomarker of certain cancers. The GFETs were characterized using direct current (DC) 4-probe electrical resistance (4-PER) measurements, which demonstrated a limit of detection of the biosensors to be ∼ 300 fg/mL (4 fM). Comparison with back-gated Dirac voltage shifts with varying concentration of Clusterin show 4-PER measurements to be more accurate, at present, and point to a requirement for further optimisation of the fabrication processes for our next generation of GFET sensors. Thus, we have successfully fabricated a promising set of GFET biosensors for the detection of Clusterin protein biomarker. The developed GFET biosensors are entirely generic and also have the potential to be applied to a variety of other disease detection applications such as Parkinson’s, cancer, and cardiovascular.
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