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1.
Talanta ; 225: 122021, 2021 Apr 01.
Article in English | MEDLINE | ID: mdl-33592751

ABSTRACT

Frequent on-line and automated monitoring of multiple protein biomarkers level secreted in the culture media during tissue growth is essential for the successful development of Tissue Engineering and Regenerative Medicine (TERM) products. Here, we present a low-cost, rapid, reliable, and integrable anion-exchange membrane-(AEM) based multiplexed sensing platform for this application. Unlike the gold-standard manual ELISA test, incubation/wash steps are optimized for each target and precisely metered in microfluidic chips to enhance selectivity. Unlike optical detection and unreliable visual detection for the ELISA test, which require standardization for every usage, the AEM ion current signal also offers robustness, endowed by the pH and ionic strength control capability of the ion-selective membrane, such that a universal standard curve can be used to calibrate all runs. The electrical signal is enhanced by highly charged silica nanoparticle reporters, which also act as hydrodynamic shear amplifiers to enhance selectivity during wash. This AEM-based sensing platform is tested with vascular protein biomarkers, Endothelin-1 (ET-1), Angiogenin (ANG) and Placental Growth Factor (PlGF). The limit of detection and three-decade dynamic range are comparable to ELISA assay but with a significantly reduced assay time of 1 h vs 7 h, due to the elimination of calibration and blocking steps. Optimized protocol for each target renders the detection highly reliable with more than 98% confidence. The multiplexed detection capability of the platform is also demonstrated by simultaneous detection of ET-1, ANG and PlGF in 40 µl of the vascular endothelial cell culture supernatants using three-membrane AEM sensor and the performance is validated against ELISA.


Subject(s)
Hydrodynamics , Silicon Dioxide , Biomarkers , Enzyme-Linked Immunosorbent Assay , Placenta Growth Factor
2.
J Phys Chem B ; 125(7): 1906-1915, 2021 02 25.
Article in English | MEDLINE | ID: mdl-33410691

ABSTRACT

The presence of a small number (∼1000) of charged nanoparticles or macromolecules on the surface of an oppositely charged perm-selective membrane is shown to sensitively gate the ionic current through the membrane at a particular voltage, thus producing a voltage signal much larger than thermal noise. We show that, at sufficiently high voltages, surface vortices appear on the membrane surface and sustain an ion-depleted boundary layer that controls the diffusion length and ion current. An asymmetric vortex bifurcation occurs beyond a critical voltage to reduce the diffusion length and the differential resistance by half. Surface nanoparticles and molecules only affect this transition voltage in the membrane I-V curve. It is shown to shift by 2 ln10 (RT/F) ∼ 0.12 V for every decade increase in bulk target concentration, independent of sensor dimension and target/probe pair. Such universal features of the surface charge-sensitive nonlinear and nonequilibrium conductance allow us to develop very robust (a 2-3 decade dynamic range for highly heterogeneous samples with built-in control) yet sensitive (subpicomolar) and selective biosensors for highly charged molecules like nucleic acids and endotoxins-and for proteins with charged nanoparticle reporters.


Subject(s)
Biosensing Techniques , Nanoparticles , Nucleic Acids , Ion Transport
3.
Commun Biol ; 2: 189, 2019.
Article in English | MEDLINE | ID: mdl-31123713

ABSTRACT

Extracellular vesicles (EV) containing microRNAs (miRNAs) have tremendous potential as biomarkers for the early detection of disease. Here, we present a simple and rapid PCR-free integrated microfluidics platform capable of absolute quantification (<10% uncertainty) of both free-floating miRNAs and EV-miRNAs in plasma with 1 pM detection sensitivity. The assay time is only 30 minutes as opposed to 13 h and requires only ~20 µL of sample as oppose to 1 mL for conventional RT-qPCR techniques. The platform integrates a surface acoustic wave (SAW) EV lysing microfluidic chip with a concentration and sensing microfluidic chip incorporating an electrokinetic membrane sensor that is based on non-equilibrium ionic currents. Unlike conventional RT-qPCR methods, this technology does not require EV extraction, RNA purification, reverse transcription, or amplification. This platform can be easily extended for other RNA and DNA targets of interest, thus providing a viable screening tool for early disease diagnosis, prognosis, and monitoring of therapeutic response.


Subject(s)
Extracellular Vesicles/chemistry , Lab-On-A-Chip Devices , MicroRNAs/blood , Animals , Biomarkers/blood , Equipment Design , Humans , Liver Neoplasms/blood , Liver Neoplasms/genetics , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , MicroRNAs/genetics , Real-Time Polymerase Chain Reaction
4.
Electrophoresis ; 2018 Feb 27.
Article in English | MEDLINE | ID: mdl-29484678

ABSTRACT

Exosomes carry microRNA biomarkers, occur in higher abundance in cancerous patients than in healthy ones, and because they are present in most biofluids, including blood and urine, these can be obtained noninvasively. Standard laboratory techniques to isolate exosomes are expensive, time consuming, provide poor purity, and recover on the order of 25% of the available exosomes. We present a new microfluidic technique to simultaneously isolate exosomes and preconcentrate them by electrophoresis using a high transverse local electric field generated by ion-depleting ion-selective membrane. We use pressure-driven flow to deliver an exosome sample to a microfluidic chip such that the transverse electric field forces them out of the cross flow and into an agarose gel which filters out unwanted cellular debris while the ion-selective membrane concentrates the exosomes through an enrichment effect. We efficiently isolated exosomes from 1× PBS buffer, cell culture media, and blood serum. Using flow rates from 150 to 200 µL/h and field strengths of 100 V/cm, we consistently captured between 60 and 80% of exosomes from buffer, cell culture media, and blood serum as confirmed by both fluorescence spectroscopy and nanoparticle tracking analysis. Our microfluidic chip maintained this recovery rate for more than 20 min with a concentration factor of 15 for 10 min of isolation.

5.
Anal Methods ; 9(28): 4112-4134, 2017.
Article in English | MEDLINE | ID: mdl-29151901

ABSTRACT

Surface acoustic waves (SAWs), are electro-mechanical waves that form on the surface of piezoelectric crystals. Because they are easy to construct and operate, SAW devices have proven to be versatile and powerful platforms for either direct chemical sensing or for upstream microfluidic processing and sample preparation. This review summarizes recent advances in the development of SAW devices for chemical sensing and analysis. The use of SAW techniques for chemical detection in both gaseous and liquid media is discussed, as well as recent fabrication advances that are pointing the way for the next generation of SAW sensors. Similarly, applications and progress in using SAW devices as microfluidic platforms are covered, ranging from atomization and mixing to new approaches to lysing and cell adhesion studies. Finally, potential new directions and perspectives on the field as it moves forward are offered, with a specific focus on potential strategies for making SAW technologies for bioanalytical applications.

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