Viral load measurements are an essential tool for the long-term clinical care of human immunodeficiency virus （HIV）-positive individuals. The gold standards in viral load instrumentation, however, are still too limi...Viral load measurements are an essential tool for the long-term clinical care of human immunodeficiency virus （HIV）-positive individuals. The gold standards in viral load instrumentation, however, are still too limited by their size, cost, and sophisticated operation for these measurements to be ubiquitous in remote settings with poor healthcare infrastructure, including parts of the world that are disproportionately affected by HIV infection. The challenge of developing a point-of-care platform capable of making viral load more accessible has been frequently approached but no solution has yet emerged that meets the practical requirements of low cost, portability, and ease-of-use. In this paper, we perform reverse-transcription loop-mediated isothermal amplification （RT-LAMP） on minimally processed HIV-spiked whole blood samples with a microfluidic and silicon microchip platform, and perform fluorescence measurements with a consumer smartphone. Our integrated assay shows amplification from as few as three viruses in a - 60 nL RT- LAMP droplet, corresponding to a whole blood concentration of 670 viruses per μL of whole blood. The technology contains greater power in a digital RT-LAMP approach that could be scaled up for the determination of viral load from a finger prick of blood in the clinical care of HIV-positive individuals. We demonstrate that all aspects of this viral load approach, from a drop of blood to imaging the RT-LAMP reaction, are compatible with lab-on-a-chip components and mobile instrumentation.展开更多
Focal adhesions are critical cell membrane components that regulate adhesion and migration and have cluster dimensions that correlate closely with adhesion engagement and migration speed.We utilized a label-free appro...Focal adhesions are critical cell membrane components that regulate adhesion and migration and have cluster dimensions that correlate closely with adhesion engagement and migration speed.We utilized a label-free approach for dynamic,long-term,quantitative imaging of cell–surface interactions called photonic resonator outcoupler microscopy(PROM)in which membrane-associated protein aggregates outcoupled photons from the resonant evanescent field of a photonic crystal biosensor,resulting in a highly localized reduction of the reflected light intensity.By mapping the changes in the resonant reflected peak intensity from the biosensor surface,we demonstrate the ability of PROM to detect focal adhesion dimensions.Similar spatial distributions can be observed between PROM images and fluorescence-labeled images of focal adhesion areas in dental epithelial stem cells.In particular,we demonstrate that cell–surface contacts and focal adhesion formation can be imaged by two orthogonal label-free modalities in PROM simultaneously,providing a general-purpose tool for kinetic,high axial-resolution monitoring of cell interactions with basement membranes.展开更多
基金supported by funding from the National Institutes of Health (NIH) Exploratory/Developmental Grant (R21) (AI106024)supported by a Ruth L.Kirschstein National Research Service Award for Individual Pred octoral MD/PhD and Other Dual Doctoral Degree Fellows (F30) (AI109825)
文摘Viral load measurements are an essential tool for the long-term clinical care of human immunodeficiency virus （HIV）-positive individuals. The gold standards in viral load instrumentation, however, are still too limited by their size, cost, and sophisticated operation for these measurements to be ubiquitous in remote settings with poor healthcare infrastructure, including parts of the world that are disproportionately affected by HIV infection. The challenge of developing a point-of-care platform capable of making viral load more accessible has been frequently approached but no solution has yet emerged that meets the practical requirements of low cost, portability, and ease-of-use. In this paper, we perform reverse-transcription loop-mediated isothermal amplification （RT-LAMP） on minimally processed HIV-spiked whole blood samples with a microfluidic and silicon microchip platform, and perform fluorescence measurements with a consumer smartphone. Our integrated assay shows amplification from as few as three viruses in a - 60 nL RT- LAMP droplet, corresponding to a whole blood concentration of 670 viruses per μL of whole blood. The technology contains greater power in a digital RT-LAMP approach that could be scaled up for the determination of viral load from a finger prick of blood in the clinical care of HIV-positive individuals. We demonstrate that all aspects of this viral load approach, from a drop of blood to imaging the RT-LAMP reaction, are compatible with lab-on-a-chip components and mobile instrumentation.
基金supported by the National Science Foundation(NSF)Grant CBET 11-32301National Institutes of Health(NIH)R01 DK099528 and NIH R21 EB018481。
文摘Focal adhesions are critical cell membrane components that regulate adhesion and migration and have cluster dimensions that correlate closely with adhesion engagement and migration speed.We utilized a label-free approach for dynamic,long-term,quantitative imaging of cell–surface interactions called photonic resonator outcoupler microscopy(PROM)in which membrane-associated protein aggregates outcoupled photons from the resonant evanescent field of a photonic crystal biosensor,resulting in a highly localized reduction of the reflected light intensity.By mapping the changes in the resonant reflected peak intensity from the biosensor surface,we demonstrate the ability of PROM to detect focal adhesion dimensions.Similar spatial distributions can be observed between PROM images and fluorescence-labeled images of focal adhesion areas in dental epithelial stem cells.In particular,we demonstrate that cell–surface contacts and focal adhesion formation can be imaged by two orthogonal label-free modalities in PROM simultaneously,providing a general-purpose tool for kinetic,high axial-resolution monitoring of cell interactions with basement membranes.
