4 results on '"Niamat Khuda"'
Search Results
2. Building a nucleic acid nanostructure with DNA-epitope conjugates for a versatile electrochemical protein detection platform
- Author
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Asanka Gurukandure, Subramaniam Somasundaram, Amanda S. N. Kurian, Niamat Khuda, and Christopher J. Easley
- Abstract
The recent surge of effort in nucleic acid-based electrochemical (EC) sensors has been fruitful, and some have even shown real-time quantification of drugs in the blood of living animals. Yet there remains a need for more generalizable EC platforms for the detection of multiple classes of clinically relevant targets. Our group has recently reported a nucleic acid nanostructure that permits simple, economical, and generalizable EC readout of a wide range of analytes (small molecules, peptides, large proteins, or antibodies). The DNA nanostructure is built through on-electrode enzymatic ligation of three oligonucleotides for attachment, binding, and signaling. However, the signaling mechanism predominantly relies on tethered diffusion of methylene blue at the electrode surface, limiting the detection of larger proteins that have no readily available small molecule binding partners. In this study, we adapted the nanostructure sensor to quantify larger proteins in a more generic manner, through conjugating the proteins minimized antibody-binding epitope to the central DNA strand of the nanostructure (DNA-peptide conjugate). This concept was verified using creatine kinase (CK-MM), an important biomarker of muscle damage, myocardial infarction, overexertion/rhabdomyolysis, or neuromuscular disorders where clinical outcomes could be improved with rapid sensing. DNA-epitope conjugates permitted a competitive immunoassay protocol at the electrode surface for quantifying CK protein. Square-wave voltammetry (SWV) signal suppression was proportional to the amount of surface-bound antibody with a limit of detection (LOD) of 5 nM and a response time as low as 3 minutes, and displacement of antibody by native CK-MM protein analyte could also be assayed. CK was quantified from the LOD of 14 nM up to 100 nM, overlapping well with the normal (non-elevated) human clinical range of 3 37 nM, and the sensor response was validated in 98% human serum. While a need for improved DNA-epitope conjugate purification was identified, overall this approach not only allows the detection of a generic protein- or peptide-binding antibody, but it also should facilitate future quantitative EC readout of various clinically relevant protein analytes that were previously inaccessible to EC techniques.
- Published
- 2023
3. Electrochemical Sensing of the Peptide Drug Exendin-4 Using a Versatile Nucleic Acid Nanostructure
- Author
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Niamat Khuda, Subramaniam Somasundaram, and Christopher J. Easley
- Subjects
Fluid Flow and Transfer Processes ,Nucleic Acids ,Process Chemistry and Technology ,Exenatide ,Humans ,Bioengineering ,DNA ,Peptides ,Instrumentation ,Article ,Nanostructures - Abstract
Although endogenous peptides and peptide-based therapeutics are both highly relevant to human health, there are few approaches for sensitive biosensing of this class of molecules with minimized workflow. In this work, we have further expanded on the generalizability of our recently developed DNA nanostructure architecture by applying it to electrochemical (EC) peptide quantification. While DNA-small molecule conjugates were used in a prior work to make sensors for small molecule and protein analytes, here DNA-peptide conjugates were incorporated into the nanostructure at the electrode surfaces, and antibody displacement permitted rapid peptide sensing. Interestingly, multivalent DNA-peptide conjugates were found to be detrimental to the assay readout, yet these effects could be minimized by solution-phase bioconjugation. The final biosensor was validated for quantifying exendin-4 (4.2 kDa)─a human glucagon-like peptide-1 receptor agonist important in diabetes therapy─for the first time using EC methods with minimal workflow. The sensor was functional in 98% human serum, and the low nanomolar assay range lies between the injected dose concentration and the therapeutic range, boding well for future applications in therapeutic drug monitoring.
- Published
- 2022
4. Non-Faradaic Current Suppression in DNA Based Electrochemical Assays with a Differential Potentiostat
- Author
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Christopher J. Easley, Mark D. Holtan, Niamat Khuda, and Subramaniam Somasundaram
- Subjects
Bioanalysis ,Chemistry ,business.industry ,DNA ,Electrochemical Techniques ,Chronoamperometry ,Electrochemistry ,Electric Capacitance ,Capacitance ,Potentiostat ,Article ,Analytical Chemistry ,Methylene Blue ,Electrode ,Optoelectronics ,Humans ,Gold ,Cyclic voltammetry ,business ,Voltammetry ,Electrodes - Abstract
One of the key factors limiting sensitivity in many electrochemical assays is the non-faradaic or capacitive current. This is particularly true in modern assay systems based on DNA monolayers at gold electrode surfaces, which have shown great promise for bioanalysis in complex milieu such as whole blood or serum. While various changes in analytical parameters, redox reporter molecules, DNA structures, probe coverage, and electrode surface area have been shown useful, background reduction by hardware subtraction has not yet been explored for these assays. Here, we introduce new electrochemistry hardware that considerably suppresses non-faradaic currents through real-time analog subtraction during current-to-voltage conversion in the potentiostat. This differential potentiostat (DiffStat) configuration is shown to suppress or remove capacitance currents in chronoamperometry, cyclic voltammetry, and square-wave voltammetry measurements applied to nucleic acid hybridization assays at the electrode surface. The DiffStat makes larger electrodes and higher sensitivity settings accessible to the user, providing order-of-magnitude improvements in sensitivity, and it also significantly simplifies data processing to extract faradaic currents in square-wave voltammetry (SWV). Since two working electrodes are used for differential measurements, unique arrangements are introduced such as converting signal-OFF assays to signal-ON assays, or background drift correction in 50 % human serum. Overall, this new potentiostat design should be helpful not only in improving the sensitivity of most electrochemical assays, but it should also better support adaptation of assays to the point-of-care by circumventing complex data processing.
- Published
- 2019
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