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1.
ACS Sens ; 8(6): 2255-2262, 2023 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-37276452

RESUMO

Loop-mediated isothermal amplification (LAMP) is a nucleic acid amplification technique that has been widely used for the detection of pathogens in many organisms. Current LAMP-based sensors usually require the LAMP products to be labeled in order for them to be detected. Here, we present a novel label-free LAMP chip, which consists of a nanopore thin-film sensor embedded inside a LAMP reaction chamber. A fraction of LAMP primers is immobilized on the sensor surface, allowing the LAMP products to be synthesized and bound to the sensor surface via immobilized primers. After the LAMP reaction components are removed from the reaction chamber, the amplified LAMP products bound to the sensor surface give rise to significantly increased transducing signals, which can be measured by a portable optical spectrometer through an optical fiber probe. As a demonstration, we used the LAMP chip to detect the causal agent of late blight, Phytophthora infestans, which is one of the most devastating plant pathogens and poses a major threat to sustainable crop production worldwide. We show that this chip can detect as low as 1 fg/µL of P. infestans DNA in 30 min, which corresponds to an attomolar level of 1.6 × 10-6 attomole/µL and is at least 10 times more sensitive than the currently available methods. This label-free sensing technology holds great promise to open up a new avenue for ultrasensitive, highly specific, rapid, and cost-effective point-of-care diagnostics of plant, animal, human, and foodborne pathogens.


Assuntos
Técnicas de Amplificação de Ácido Nucleico , Animais , Humanos , Técnicas de Amplificação de Ácido Nucleico/métodos , Primers do DNA/genética
2.
Lab Chip ; 23(6): 1649-1663, 2023 03 14.
Artigo em Inglês | MEDLINE | ID: mdl-36751868

RESUMO

To improve our understanding of how the central nervous system functions in health and disease, we report the development of an integrated chip for studying the effects of the neurotransmitters dopamine and serotonin on adult rat hippocampal progenitor cell (AHPC) neurospheroids. This chip allows dopamine or serotonin located in one chamber to diffuse to AHPC neurospheroids cultured in an adjacent chamber through a built-in diffusion barrier created by an array of intentionally misaligned micropillars. The gaps among the micropillars are filled with porous poly(ethylene glycol) (PEG) gel to tune the permeability of the diffusion barrier. An electrochemical sensor is also integrated within the chamber where the neurospheroids can be cultured, thereby allowing monitoring of the concentrations of dopamine or serotonin. Experiments show that concentrations of the neurotransmitters inside the neurospheroid chamber can be increased over a period of several hours to over 10 days by controlling the compositions of the PEG gel inside the diffusion barrier. The AHPC neurospheroids cultured in the chip remain highly viable following dopamine or serotonin treatment. Cell proliferation and neuronal differentiation have also been observed following treatment, revealing that the AHPC neurospheroids are a valuable in vitro brain model for neurogenesis research. Finally, we show that by tuning the permeability of diffusion barrier, we can block transfer of Escherichia coli cells across the diffusion barrier, while allowing dopamine or serotonin to pass through. These results suggest the feasibility of using the chip to better understand the interactions between microbiota and brain via the gut-brain axis.


Assuntos
Dopamina , Microfluídica , Ratos , Animais , Serotonina , Encéfalo , Neurotransmissores
3.
Lab Chip ; 21(16): 3128-3136, 2021 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-34180491

RESUMO

Integrins are key players in platelet adhesion and aggregation. Integrin molecular tensions, the forces transmitted by integrin molecules, are regulated by both mechanical and biochemical cues, and the outside-in and inside-out signaling has been extensively studied. While the mechanical properties of platelets at static status have been studied by atomic force microscopy, traction force microscopy and tension sensors, the biomechanical properties of flowing platelets remain elusive. Herein, we report microfluidic chips grafted with integrin tension sensors for microfluidic-force mapping in platelets. Specifically, the process of integrin αIIbß3 mediating tension transmission and platelet adhesion under low flow rates has been obtained, and the process of platelet clustering at post-stenotic regions has been demonstrated. We found that flowing shear force can postpone the integrin-mediated tension transmission and platelet adhesion. We further evaluated the effect of Y-27632, a ROCK inhibitor that has been proven to reduce integrin-mediated platelet adhesion, at a series of concentrations and demonstrated that microfluidic chips with integrin tension sensors are sensitive to the concentration-dependent effects of Y-27632. Given their low cost and scalable throughput, these chips are ideal technical platforms for biological studies of platelets at flowing status and for platelet inhibitor or potential antiplatelet drug screening.


Assuntos
Plaquetas , Integrinas , Microfluídica , Quinases Associadas a rho/antagonistas & inibidores , Animais , Cães , Adesividade Plaquetária , Estresse Mecânico
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