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Articles by Vivienne N. Luk in JoVE
JoVE General
स्वचालित प्रोटिओमिक प्रसंस्करण के लिए डिजिटल Microfluidics
1Department of Chemistry, University of Toronto, 2Donnelly Centre for Cellular and Biomolecular Research, 3Institute for Biomaterials and Biomedical Engineering, University of Toronto
बिजली क्षेत्र के अनुप्रयोग द्वारा इलेक्ट्रोड की एक सरणी पर - डिजिटल Microfluidics एक तकनीक असतत बूंदों (एमएल ~ nl) के हेरफेर की विशेषता है. यह अच्छी तरह से बाहर तेजी से, अनुक्रमिक, छोटी स्वचालित जैव रासायनिक assays ले जाने के लिए अनुकूल है. यहाँ, हम एक कई प्रोटिओमिक प्रसंस्करण कदम को स्वचालित करने में सक्षम मंच की रिपोर्ट.
Other articles by Vivienne N. Luk on PubMed
Pluronic Additives: a Solution to Sticky Problems in Digital Microfluidics
Digital microfluidics (DMF) is a promising technique for carrying out miniaturized, automated biochemical assays in which discrete droplets of reagents are actuated on the surface of an array of electrodes. A limitation for DMF is nonspecific protein adsorption to device surfaces, which interferes with assay fidelity and can cause droplets to become unmovable. Here, we report the results of a quantitative analysis of protein adsorption on DMF devices by means of confocal microscopy and secondary ion mass spectrometry. This study led us to a simple and effective method for limiting the extent of protein adsorption: the use of low concentrations of Pluronic F127 as a solution additive. This strategy has a transformative effect on digital microfluidics, facilitating the actuation of droplets containing greater than 1000-fold higher protein concentrations than is possible without the additive. To illustrate the benefits of this new method, we implemented a DMF-driven protein digest assay using large concentrations (1 mg/mL) of protein-substrate. The use of Pluronic additives solves a sticky problem in DMF, which greatly expands the range of applications that are compatible with this promising technology.
A World-to-chip Interface for Digital Microfluidics
Digital microfluidics (DMF) is a fluid handling technique that enables manipulation of discrete droplets on an array of electrodes. There is considerable enthusiasm for this method because of the potential for array-based screening applications. A limitation for DMF is nonspecific adsorption of reagents to device surfaces. If a given device is used to actuate multiple reagents, this phenomenon can cause undesirable cross-contamination. A second limitation for DMF (and all other microfluidic systems) is the "world-to-chip" interface; it is notoriously difficult to deliver reagents and samples to such systems without compromising the oft-hyped advantages of rapid analyses and reduced reagent consumption. We introduce a new strategy for digital microfluidics, in which a removable plastic "skin" is used to (a) eliminate cross-contamination and (b) bridge the world-to-chip interface. We demonstrated the utility of this format by implementing on-chip protein digestion on immobilized enzyme depots. This new method has the potential to transform DMF from being a curiosity for aficionados into a technology that is useful for biochemical applications at large.
A Digital Microfluidic Approach to Proteomic Sample Processing
A common characteristic for proteomic analyses is the need for extensive biochemical processing. Digital microfluidics (DMF), a technique characterized by the manipulation of discrete microdroplets (100 nL-10 microL) on an open array of electrodes, is a good match for carrying out rapid, automated solution-phase reactions. Here, we report a DMF-based method integrating several common processing steps in proteomics, including reduction, alkylation, and enzymatic digestion. Fluorogenic assays were used to quantitatively evaluate the kinetics and reproducibility of each reaction step, and MALDI-MS was used for qualitative confirmation. The method is fast, facile, and reproducible, and thus has the potential to be a useful new tool in proteomics.
