Questo lavoro è stato finanziato dal Fonds de Recherche sur la Nature et les Technologies de Quebec, in Canada la Fondazione per l&#39;Innovazione e la Canadian Institutes of Health Research (CIHR).

1. Microfabbricazione della testa della sonda (processo non mostrato nel video) <ol><li> A Si 2 wafer, quattro pollici di diametro, 525 micron di spessore, con uno strato di 1 micron di spessore SiO2 termico è spincoated con photoresist (PR) per 45 s a 4000 giri. </li><li> Il wafer è precotti a 110C per 50 s, ed esposta attraverso una maschera con tutti gli elementi (porte e microcanali) per 5 s, sviluppato e sciacquato con DI. </li><li> La scoperta è inciso SiO2 via in una tamponata 01:07 acido fluoridrico (BHF) una soluzione ≈ 15 min (dewetting del substrato in cui SiO2 è stato inciso indica il completamento della etch). Un plasma O2 o acetone viene utilizzato in cenere o nastri il restante PR. </li><li> Un secondo strato PR è spin-rivestito a 1500 rpm per 45 s, ottenendo un overlayer spessore di ≈ 10 micron [31]. Il modello SiO2 sotto questo strato di PR è ancora visibile e viene utilizzato per allineare il wafer con una seconda maschera con le uniche porte. </li><li> Dopo l&#39;esposizione e lo sviluppo del PR, il wafer viene sciacquato, asciugato e postbaked a 95C per 20 min. </li><li> Il wafer di silicio è fissato su un wafer di supporto con fuso cera bianca per proteggere il mandrino. </li><li> Un plasma ad accoppiamento induttivo (ICP) DRIE è utilizzato per trasferire il PR ed embedded SiO2 modelli in topografia wafer in un processo in tre fasi: <ol><li> DRIE fare ≈ 500μmdeep porte in Si (modello definita dalla spessa PR). </li><li> Senza scaricare il wafer dalla macchina DRIE, il PR è incenerito con un plasma. </li><li> Il modello esposto SiO2 agisce come maschera per un secondo processo a secco etch, la creazione di 50 canali micron profondo, e aprendo il ripieno e le porte di ventilazione attraverso il wafer. Dopo lo scarico, il wafer supporto è staccata sotto un getto di acqua calda. Il wafer microlavorati viene pulito con acetone, etanolo e DI. </li></ol></li><li> Singoli chip MFP sono a dadini. </li><li> Un blocco di interfaccia PDMS è fabbricata la fusione in uno micromould composto da due strutturato poli (metilmetacrilato) (PMMA) elementi, una piastra in acciaio lucido che formano la parte inferiore, e due capillari (ciascuno inserito in una delle due vias accesso buchi nel lamiera di acciaio) che serve come segnaposto per i fori di collegamento fluidica. Il PDMS è curato in forno a 60 ° C per almeno 1 h. </li><li> Il blocco PDMS è legato ad un dadini chip di silicio MFP, attivando entrambe le parti in aria plasma a 1 mbar per 24 s a 230W, e unendo le due cose insieme con un home-made aiuto per l&#39;allineamento meccanico. </li><li> L&#39;assemblea è lasciato al bond in un forno a 60 ° C per un minimo di 1 ora </li></ol> 2. Assemblea della MFP <ol><li> Tenuta di gas siringhe di vetro sono riempiti con i reagenti appropriati con siringhe e aghi di plastica per garantire che non siano presenti bolle d&#39;aria. Tipicamente, si usa una siringa da 1 -10 microlitro per iniezione, e una siringa con un volume di 5-10 volte più grande per l&#39;aspirazione. </li><li> Le siringhe sono collegati a tubi capillari utilizzando raccordi Nanotight a basso volume morto. </li><li> Capillari sono riempiti e controllati per bolle sotto il microscopio. </li><li> Il chip MFP è pre-riempite con soluzione tampone per evitare cattura di bolla quando si collegano i capillari. </li><li> I capillari sono collegati al raccordo PDMS nella testa della sonda </li></ol> 3. Set-up della MFP <ol><li> La testa della sonda è bloccato nel supporto sonda e montato sulla sonda stazione in cima ad un microscopio invertito </li><li> Le siringhe sono posti in alta precisione pompe siringa. </li><li> Il substrato, come ad esempio un vetrino, viene inserito in una porta fatta in casa che viene apposta sul palco microscopio. </li><li> Il parallelismo della mesa del MFP e il substrato è regolato con un paio di goniometri osservando gli anelli di Newton (frange di interferenza) che appaiono quando la stampante MFP è portato a contatto con il substrato. Il punto di contatto e la frequenza degli anelli servono come indicazione del tilt. Quando la stampante MFP è allineato con la superficie, un anello di interferenza singolo si estende su tutta la superficie. Questa misura serve anche per calibrare la separazione tra MFP e substrato. </li><li> Il divario tra il MFP e il substrato è un fattore critico per i processi di patterning superficie. Perché il substrato è elaborato dalla scansione al di sotto del MFP, l&#39;allineamento orizzontale deve essere regolato con precisione micrometrica ed è realizzato utilizzando un supporto a tre punti formato da tre viti micrometriche. </li></ol> 4. Funzionamento del MFP <ol><li> Erogazione è controllata via software LabView. Funzionamento del dispositivo viene visualizzato ad occhio e utilizzando una telecamera CCD. L&#39;iniezione: rapporto aspirazione varia 1:03-1:10, a seconda della diffusività del reagente con il tampone circostante e il modello desiderato flusso geometrico. </li><li> Per controllare il corretto funzionamento della siringa di aspirazione e la presenza di bolle, prima iniettare liquido con la siringa di aspirazione prima di iniziare l&#39;aspirazione corretta.</li><li> Inizio iniezione di liquido e il flusso di monitorare e confinamento di perline o di colorante tracciante fluorescente. </li><li> Utilizzare la sonda per la particolare applicazione, cioè la scansione attraverso la superficie per la lavorazione per la deposizione, di attacco o macchie di superficie o di cellule. </li></ol>

<ol>
	<li>Juncker, D., Schmid, H., Delamarche, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Nature Materials. 4 (8), 622-622 (2005).</li></ol>

La sonda microfluidica (MFP) è versatile perché è (i) mobili, (ii) adattabile per essere utilizzato con diversi tipi di reagenti e di substrato e può (iii) essere utilizzato su grandi aree. Bolle indesiderate può portare alla interruzione del flusso per evitare bolle, tutti i componenti devono essere pieni di liquidi prima del montaggio. Il divario tra la sonda e la superficie è solo di pochi micrometri, ma il mesa è diverse centinaia di micron di larghezza, e distanze nel range di ce...

<table><tbody><tr><td>microfluidic connectors</td><td>Upchurch Scientific</td><td>Micro- and Nano-tight fittings and sleeves</td><td></td></tr><tr><td>2-component manual dispenser</td><td>Conprotec Inc.</td><td>DM400</td><td>To dispense and mix PDMS mixture</td></tr><tr><td>LabVIEW</td><td>National Instruments</td><td>Version 8.0</td><td></td></tr><tr><td>Mechanical Convection Oven</td><td>VWR international</td><td>1330FM</td><td></td></tr><tr><td>Glass syringes</td><td>Hamilton Co</td><td></td><td></td></tr><tr><td>Capillary tubing</td><td>Polymicro Technologies</td><td></td><td></td></tr><tr><td>Plasma Chamber</td><td>Tegal Corporation</td><td>Plasmaline 415</td><td></td></tr><tr><td>Inverted Microscope</td><td>Nikon Instruments</td><td>TE2000-E</td><td></td></tr><tr><td>Syringe pumps</td><td>Cetoni</td><td>neMESYS</td><td></td></tr><tr><td>Sylgard 184</td><td>Ellsworth Adhesives</td><td>184 Sil Elast Kit</td><td></td></tr><tr><td>Camera</td><td>Photometrics</td><td>QuantEM 512SC</td><td></td></tr><tr><td>Microscope stage</td><td></td><td></td><td></td></tr><tr><td>Microfluidic probe holder goniometers</td><td>Melles Griot</td><td>07GON504</td><td></td></tr><tr><td>Linear stage</td><td>Applied Scientific Instrumentation</td><td>LS-50</td><td>For z-control of the MFP</td></tr><tr><td>Manual linear stage</td><td>Newport Corp.</td><td>443-4 Series</td><td>For x- and y- axis control of the MFP</td></tr><tr><td>Microscope stage</td><td>Applied Scientific Instrumentation</td><td>PZ-2000</td><td>With x-, y- and z- control</td></tr></tbody></table>

the microfluidic probe: operation and use for localized surface processing

Dispositivi microfluidici permettono test deve essere eseguito utilizzando piccole quantità di campione e sono stati recentemente utilizzati per controllare il microambiente delle cellule. Microfluidica è comunemente associato con microcanali chiuse che limitano il loro uso per campioni che possono essere introdotte, e colto nel caso delle cellule, all&#39;interno di un volume confinato. D&#39;altra parte, micropipetting sistema sono stati utilizzati per perfusione a livello locale le cellule e le superfici, in particolare con push-pull configurazioni in cui si agisce pipetta come sorgente e l&#39;altra come affondare, ma il confinamento del flusso è difficile in tre dimensioni. Inoltre, pipette sono fragili e difficili da posizione e, quindi, sono utilizzati in configurazione statica solo. La microfluidica sonda (MFP) elude i vincoli imposti dalla costruzione di chiuse e canali microfluidica invece che racchiude il campione nel sistema microfluidica, il flusso microfluidica può essere consegnato direttamente sul campione, e la scansione attraverso il campione, usando la MFP. . L&#39;iniezione e aperture di aspirazione si trovano a poche decine di micrometri l&#39;uno dall&#39;altro in modo che un microjet iniettato il divario è confinato dalle forze idrodinamiche del liquido circostante e interamente aspirato nuovamente dentro l&#39;altra apertura. La microjet può essere lavato tutta la superficie del substrato e fornisce uno strumento preciso per la deposizione localizzata / fornitura di reagenti che possono essere utilizzate su vaste aree mediante la scansione di sonda su tutta la superficie. In questo video vi presentiamo la microfluidica sonda 1 (MFP). Vi spieghiamo in dettaglio come assemblare il MFP, montarlo in cima a un microscopio invertito, e allinearlo rispetto alla superficie del substrato e, infine, mostrano come usarlo per elaborare una superficie substrato immerso in un buffer.

Watch this Scientific Journal Video about The Microfluidic Probe: Operation and Use for Localized Surface Processing at JoVE.com

The Microfluidic Probe: Operation and Use for Localized Surface Processing

In questo video vi presentiamo la sonda microfluidica<sup&gt; 1</sup&gt; (MFP). Vi spieghiamo in dettaglio come assemblare il MFP, montarlo in cima a un microscopio invertito, e allinearlo rispetto alla superficie del substrato e, infine, mostrano come usarlo per elaborare una superficie substrato immerso in una soluzione tampone.

La sonda Microfluidic: funzionamento e utilizzo per l&#39;elaborazione delle superfici localizzate

Microfluidic devices allow assays to be performed using minute amounts of sample and have recently been used to control the microenvironment of cells. ...

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Research

JoVE Journal

Biology

8.8K Views.  McGill University. In questo video vi presentiamo la sonda microfluidica 1 (MFP). Vi spieghiamo in dettaglio come assemblare il MFP, montarlo in cima a un microscopio invertito, e allinearlo rispetto alla superficie del substrato e, infine, mostrano come usarlo per elaborare una superficie substrato immerso in una soluzione tampone.

Video: The Microfluidic Probe: Operation and Use for Localized Surface Processing

Fluorescence detection methods for microfluidic droplet platforms

The development of microfluidic platforms for performing chemistry and biology has in large part been driven by a range of potential benefits that accompany system miniaturisation. Advantages include the ability to efficiently process nano- to femoto- liter volumes of sample, facile integration of functional components, an intrinsic predisposition towards large-scale multiplexing, enhanced analytical throughput, improved control and reduced instrumental footprints.1
In recent years much interest has focussed on the development of droplet-based (or segmented flow) microfluidic systems and their potential as platforms in high-throughput experimentation.2-4 Here water-in-oil emulsions are made to spontaneously form in microfluidic channels as a result of capillary instabilities between the two immiscible phases. Importantly, microdroplets of precisely defined volumes and compositions can be generated at frequencies of several kHz. Furthermore, by encapsulating reagents of interest within isolated compartments separated by a continuous immiscible phase, both sample cross-talk and dispersion (diffusion- and Taylor-based) can be eliminated, which leads to minimal cross-contamination and the ability to time analytical processes with great accuracy. Additionally, since there is no contact between the contents of the droplets and the channel walls (which are wetted by the continuous phase) absorption and loss of reagents on the channel walls is prevented.
Once droplets of this kind have been generated and processed, it is necessary to extract the required analytical information. In this respect the detection method of choice should be rapid, provide high-sensitivity and low limits of detection, be applicable to a range of molecular species, be non-destructive and be able to be integrated with microfluidic devices in a facile manner. To address this need we have developed a suite of experimental tools and protocols that enable the extraction of large amounts of photophysical information from small-volume environments, and are applicable to the analysis of a wide range of physical, chemical and biological parameters. Herein two examples of these methods are presented and applied to the detection of single cells and the mapping of mixing processes inside picoliter-volume droplets. We report the entire experimental process including microfluidic chip fabrication, the optical setup and the process of droplet generation and detection.

Droplet-based microfluidic platforms are promising candidates for high throughput experimentation since they are able to generate picoliter, self-compartmentalized vessels inexpensively at kHz rates. Through integration with fast, sensitive and high resolution fluorescence spectroscopic methods, the large amounts of information generated within these systems can be efficiently extracted, harnessed and utilized.

Metodi di rilevamento a fluorescenza per piattaforme goccia microfluidica

The development of microfluidic platforms for performing chemistry and biology has in large part been driven by a range of potential benefits that ...

Ricerca

Bioingegneria

Taking Advantage of Reduced Droplet-surface Interaction to Optimize Transport of Bioanalytes in Digital Microfluidics

Digital microfluidics (DMF), a technique for manipulation of droplets, is a promising alternative for the development of &#8220;lab-on-a-chip&#8221; platforms. Often, droplet motion relies on the wetting of a surface, directly associated with the application of an electric field; surface interactions, however, make motion dependent on droplet contents, limiting the breadth of applications of the technique.
Some alternatives have been presented to minimize this dependence. However, they rely on the addition of extra chemical species to the droplet or its surroundings, which could potentially interact with droplet moieties. Addressing this challenge, our group recently developed Field-DW devices to allow the transport of cells and proteins in DMF, without extra additives.
Here, the protocol for device fabrication and operation is provided, including the electronic interface for motion control. We also continue the studies with the devices, showing that multicellular, relatively large, model organisms can also be transported, arguably unaffected by the electric fields required for device operation.

The protocol for fabrication and operation of field dewetting devices (Field-DW) is described, as well as the preliminary studies of the effects of electric fields on droplet contents.

Approfittando di ridotta interazione Droplet-superficie per ottimizzare il trasporto di Bioanalytes in microfluidica digitale

Digital microfluidics (DMF), a technique for manipulation of droplets, is a promising alternative for the development of &#8220;lab-on-a-chip&#8221; ...

Ingegneria

A Microfluidic Chip for the Versatile Chemical Analysis of Single Cells

We present a microfluidic device that enables the quantitative determination of intracellular biomolecules in multiple single cells in parallel. For this purpose, the cells are passively trapped in the middle of a microchamber. Upon activation of the control layer, the cell is isolated from the surrounding volume in a small chamber. The surrounding volume can then be exchanged without affecting the isolated cell. However, upon short opening and closing of the chamber, the solution in the chamber can be replaced within a few hundred milliseconds. Due to the reversibility of the chambers, the cells can be exposed to different solutions sequentially in a highly controllable fashion, e.g. for incubation, washing, and finally, cell lysis. The tightly sealed microchambers enable the retention of the lysate, minimize and control the dilution after cell lysis. Since lysis and analysis occur at the same location, high sensitivity is retained because no further dilution or loss of the analytes occurs during transport. The microchamber design therefore enables the reliable and reproducible analysis of very small copy numbers of intracellular molecules (attomoles, zeptomoles) released from individual cells. Furthermore, many microchambers can be arranged in an array format, allowing the analysis of many cells at once, given that suitable optical instruments are used for monitoring. We have already used the platform for proof-of-concept studies to analyze intracellular proteins, enzymes, cofactors and second messengers in either relative or absolute quantifiable manner.

In this article we present a microfluidic chip for single cell analysis. It allows the quantification of intracellular proteins, enzymes, cofactors, and second messengers by means of fluorescent assays or immunoassays.&#160;

Un chip microfluidica per la Versatile analisi chimica di singole cellule

We present a microfluidic device that enables the quantitative determination of intracellular biomolecules in multiple single cells in parallel. For this ...

Combining Microfluidics and Microrheology to Determine Rheological Properties of Soft Matter during Repeated Phase Transitions

The microstructure of soft matter directly impacts macroscopic rheological properties and can be changed by factors including colloidal rearrangement during previous phase changes and applied shear. To determine the extent of these changes, we have developed a microfluidic device that enables repeated phase transitions induced by exchange of the surrounding fluid and microrheological characterization while limiting shear on the sample. This technique is &#181;2rheology, the combination of microfluidics and microrheology. The microfluidic device is a two-layer design with symmetric inlet streams entering a sample chamber that traps the gel sample in place during fluid exchange. Suction can be applied far away from the sample chamber to pull fluids into the sample chamber. Material rheological properties are characterized using multiple particle tracking microrheology (MPT). In MPT, fluorescent probe particles are embedded into the material and the Brownian motion of the probes is recorded using video microscopy. The movement of the particles is tracked and the mean-squared displacement (MSD) is calculated. The MSD is related to macroscopic rheological properties, using the Generalized Stokes-Einstein Relation. The phase of the material is identified by comparison to the critical relaxation exponent, determined using time-cure superposition. Measurements of a fibrous colloidal gel illustrate the utility of the technique. This gel has a delicate structure that can be irreversibly changed when shear is applied. &#181;2rheology data shows that the material repeatedly equilibrates to the same rheological properties after each phase transition, indicating that phase transitions do not play a role in microstructural changes. To determine the role of shear, samples can be sheared prior to injection into our microfluidic device. &#181;2rheology is a widely applicable technique for the characterization of soft matter enabling the determination of rheological properties of delicate microstructures in a single sample during phase transitions in response to repeated changes in the surrounding environmental conditions.

We demonstrate the fabrication and use of a microfluidic device that enables multiple particle tracking microrheology measurements to study the rheological effects of repeated phase transitions on soft matter.

Combinazione di microfluidica e Microrheology per determinare le proprietà reologiche dei materiali molli durante le transizioni di fase ripetute

The microstructure of soft matter directly impacts macroscopic rheological properties and can be changed by factors including colloidal rearrangement ...

A Microfluidic Platform for Precision Small-volume Sample Processing and Its Use to Size Separate Biological Particles with an Acoustic Microdevice

A major advantage of microfluidic devices is the ability to manipulate small sample volumes, thus reducing reagent waste and preserving precious sample. However, to achieve robust sample manipulation it is necessary to address device integration with the macroscale environment. To realize repeatable, sensitive particle separation with microfluidic devices, this protocol presents a complete automated and integrated microfluidic platform that enables precise processing of 0.15&#8211;1.5 ml samples using microfluidic devices. Important aspects of this system include modular device layout and robust fixtures resulting in reliable and flexible world to chip connections, and fully-automated fluid handling which accomplishes closed-loop sample collection, system cleaning and priming steps to ensure repeatable operation. Different microfluidic devices can be used interchangeably with this architecture. Here we incorporate an acoustofluidic device, detail its characterization, performance optimization, and demonstrate its use for size-separation of biological samples. By using real-time feedback during separation experiments, sample collection is optimized to conserve and concentrate sample. Although requiring the integration of multiple pieces of equipment, advantages of this architecture include the ability to process unknown samples with no additional system optimization, ease of device replacement, and precise, robust sample processing.

This protocol describes a system architecture for performing automated small volume (0.15&#8211;1.5 ml) particle separations using a microfluidic device, and discusses methods to optimize acoustofluidic device performance and operation.

Una piattaforma per l&#39;elaborazione di precisione Microfluidic piccola Volume del campione e il suo uso per Dimensione particelle biologiche separate con un Microdevice Acoustic

A major advantage of microfluidic devices is the ability to manipulate small sample volumes, thus reducing reagent waste and preserving precious sample. ...

Digital Microfluidics for Automated Proteomic Processing

Clinical proteomics has emerged as an important new discipline, promising the discovery of biomarkers that will be useful for early diagnosis and prognosis of disease. While clinical proteomic methods vary widely, a common characteristic is the need for (i) extraction of proteins from extremely heterogeneous fluids (i.e. serum, whole blood, etc.) and (ii) extensive biochemical processing prior to analysis. Here, we report a new digital microfluidics (DMF) based method integrating several processing steps used in clinical proteomics. This includes protein extraction, resolubilization, reduction, alkylation and enzymatic digestion. Digital microfluidics is a microscale fluid-handling technique in which nanoliter-microliter sized droplets are manipulated on an open surface. Droplets are positioned on top of an array of electrodes that are coated by a dielectric layer - when an electrical potential is applied to the droplet, charges accumulate on either side of the dielectric. The charges serve as electrostatic handles that can be used to control droplet position, and by biasing a sequence of electrodes in series, droplets can be made to dispense, move, merge, mix, and split on the surface. Therefore, DMF is a natural fit for carrying rapid, sequential, multistep, miniaturized automated biochemical assays. This represents a significant advance over conventional methods (relying on manual pipetting or robots), and has the potential to be a useful new tool in clinical proteomics.
Mais J. Jebrail, Vivienne N. Luk, and Steve C. C. Shih contributed equally to this work.
Sergio L. S. Freire's current address is at the University of the Sciences in Philadelphia located at 600 South 43rd Street, Philadelphia, PA 19104.

Digital Microfluidics is a technique characterized by the manipulation of discrete droplets (~nL - mL) on an array of electrodes by the application of electrical fields. It is well-suited for carrying out rapid, sequential, miniaturized automated biochemical assays. Here, we report a platform capable of automating several proteomic processing steps.

Microfluidica digitale per trattamento automatizzato di proteomica

Clinical proteomics has emerged as an important new discipline, promising the discovery of biomarkers that will be useful for early diagnosis and ...

Biologia

Rapid Subtractive Patterning of Live Cell Layers with a Microfluidic Probe

The microfluidic probe (MFP) facilitates performing local chemistry on biological substrates by confining nanoliter volumes of liquids. Using one particular implementation of the MFP, the hierarchical hydrodynamic flow confinement (hHFC), multiple liquids are simultaneously brought in contact with a substrate. Local chemical action and liquid shaping using the hHFC, is exploited to create cell patterns by locally lysing and removing cells. By utilizing the scanning ability of the MFP, user-defined patterns of cell monolayers are created. This protocol enables rapid, real-time and spatially controlled cell patterning, which can allow selective cell-cell and cell-matrix interaction studies.

We present a protocol to perform subtractive patterning of live cell monolayers on a surface. This is achieved by local and selective lysis of adherent cells using a microfluidic probe (MFP). The cell lysate retrieved from local regions can be used for downstream analysis, enabling molecular profiling studies.

Rapid Patterning sottrattiva di strati cellulare dal vivo con una sonda Microfluidic

The microfluidic probe (MFP) facilitates performing local chemistry on biological substrates by confining nanoliter volumes of liquids. Using one ...

A Microfluidic System with Surface Patterning for Investigating Cavitation Bubble(s)&#8211;Cell Interaction and the Resultant Bioeffects at the Single-cell Level

In this manuscript, we first describe the fabrication protocol of a microfluidic chip, with gold dots and fibronectin-coated regions on the same glass substrate, that precisely controls the generation of tandem bubbles and individual cells patterned nearby with well-defined locations and shapes. We then demonstrate the generation of tandem bubbles by using two pulsed lasers illuminating a pair of gold dots with a few-microsecond time delay. We visualize the bubble-bubble interaction and jet formation by high-speed imaging and characterize the resultant flow field using particle image velocimetry (PIV). Finally, we present some applications of this technique for single cell analysis, including cell membrane poration with macromolecule uptake, localized membrane deformation determined by the displacements of attached integrin-binding beads, and intracellular calcium response from ratiometric imaging. Our results show that a fast and directional jetting flow is produced by the tandem bubble interaction, which can impose a highly localized shear stress on the surface of a cell grown in close proximity. Furthermore, different bioeffects can be induced by altering the strength of the jetting flow by adjusting the standoff distance from the cell to the tandem bubbles.

A microfluidic chip was fabricated to produce pairs of gold dots for tandem bubble generation and fibronectin-coated islands for single-cell patterning nearby. The resultant flow field was characterized by particle image velocimetry and was employed to study various bioeffects, including cell membrane poration, membrane deformation, and intracellular calcium response.

Un sistema Microfluidic con Patterning superficie per indagare cavitazione Bubble (s) Interazione -cell e gli effetti biologici risultante nel livello di singola cellula

In this manuscript, we first describe the fabrication protocol of a microfluidic chip, with gold dots and fibronectin-coated regions on the same glass ...

Computer Numerical Control Micromilling of a Microfluidic Acrylic Device with a Staggered Restriction for Magnetic Nanoparticle-Based Immunoassays

Microfluidic systems have greatly improved immunoassay techniques. However, many microfabrication techniques require specialized, expensive, or complicated equipment, making fabrication costly and incompatible with mass production, which is one of the most important preconditions for point-of-care tests (POCT) to be adopted in low-resource settings. This work describes the fabrication process of an acrylic (polymethylmethacrylate, PMMA) device for nanoparticle-conjugated enzymatic immunoassay testing using the computer numerical control (CNC) micromilling technique. The functioning of the microfluidic device is shown by performing an immunoassay to detect a commercial antibody using lysozyme as a model antigen conjugated to 100 nm magnetic nanoparticles. This device integrates a physical staggered restriction of only 5 &#181;m in height, used to capture magnetic microparticles that make up a magnetic trap by placing an external magnet. In this way, the magnetic force on the immunosupport of conjugated nanoparticles is enough to capture them and resist flow drag. This microfluidic device is particularly suitable for low-cost mass production without the loss of precision for immunoassay performance.

Microfluidics is a powerful tool for the development of diagnostic tests. However, expensive equipment and materials, as well as laborious fabrication and handling techniques, are often required. Here, we detail the fabrication protocol of an acrylic microfluidic device for magnetic micro- and nanoparticle-based immunoassays in a low-cost and simple-to-use setting.

Microfresatura a controllo numerico computerizzato di un dispositivo acrilico microfluidico con restrizione sfalsata per saggi immunologici magnetici basati su nanoparticelle

Microfluidic systems have greatly improved immunoassay techniques. However, many microfabrication techniques require specialized, expensive, or ...

Microfluidic Tools for Probing Fungal-Microbial Interactions at the Cellular Level

Filamentous fungi are successful inhabitants of soil and play a major role in soil ecosystems, such as in the decomposition of organic and inorganic matter, as well as regulation of nutrient levels. There they also find numerous opportunities to interact with a variety of other microbes such as bacteria or other fungi. Studying fungal interactions at the cellular level, however, can be challenging owing to the black box-like nature of soil. New microfluidic tools are being developed for the study of fungal interactions; two platforms designed to study bacterial-fungal and fungal-fungal interactions are highlighted. Within these microchannels, fungal-microbial interactions can be monitored in controlled physico-chemical environments at higher temporal and spatial resolution than previously possible. Application of these tools have yielded numerous novel biological insights, such as the observation of bacterial polar attachment to hyphae or revealing uncharacterised fungal-fungal antagonisms. A key feature of these methodologies regards the ease of use of this tool by non-experts, yielding highly translatable technologies for use in microbiology labs.

Owing to the opacity of soil, interactions between its constituent microbes cannot easily be visualised with cellular resolution. Here, two microfluidic tools are presented, which offer new opportunities for investigating fungal-microbial interactions. The devices are versatile and simple to use, enabling high spatiotemporal control and high-resolution imaging at the cellular level.

Strumenti microfluidici per sondare le interazioni fungine-microbiche a livello cellulare

Filamentous fungi are successful inhabitants of soil and play a major role in soil ecosystems, such as in the decomposition of organic and inorganic ...

La sonda Microfluidic: funzionamento e utilizzo per l'elaborazione delle superfici localizzate

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