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RISS 인기검색어
- 검색키워드
- 저자 : Shimshon Belkin (검색결과 3 건)
- 무료
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Aptamer-based Depletion of Small Molecular Contaminants: A Case Study Using Ochratoxin A
Emilia Schax,Maren Lönne,Thomas Scheper,Shimshon Belkin,Johanna-Gabriela Walter 한국생물공학회 2015 Biotechnology and Bioprocess Engineering Vol.20 No.6
Based on the increasing demand for detection and depletion of small molecules like mycotoxins or pesticides in food, water, or pharmaceuticals, aptamers are gaining more importance as sensitive, specific-depletion molecules. Here, we present an aptamer-based method for depletion of ochratoxin A (OTA) as a model system and show the advantages and the limitations of aptamers in the depletion of small molecular contaminants. OTA is a mycotoxin produced by various Penicillium and Aspergillus strains and is often found in grain and grain derivatives. We immobilized a well-described DNA aptamer against OTA on an agarose gel and used the column as a clean-up system. The aptamer shows a high specificity and sensitivity for OTA: Ochratoxin B, a molecule similar to OTA, was not bound by the aptamer; and a control oligonucleotide was not able to bind OTA. After optimizing the process for better economic feasibility, the column could be used for several times without loss of aptamer activity. We investigated the location of immobilized aptamer within the gel using fluorescent-labeled aptamers. Furthermore, beer samples spiked with OTA were used to investigate aptamer activity in complex samples. Using these complex samples we have observed a significant loss of aptamer activity. We have further investigated this limitation by performing microscale thermophoresis experiments to determine the KD values of the aptamer in different complex samples like beer, coffee, juice and wine. Our results indicate that the applicability of aptamers to real processes is currently restricted by the selection buffer used during its selection process (SELEX). We therefore suggest using conditions closer to those of the later application of the aptamer during future SELEX experiments.
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High-throughput prescreening of pharmaceuticals using a genome-wide bacterial bioreporter array
Elad, Tal,Seo, Ho Bin,Belkin, Shimshon,Gu, Man Bock Elsevier 2015 Biosensors & bioelectronics Vol.68 No.-
<P><B>Abstract</B></P> <P>We assessed the applicability of multi-strain bacterial bioreporter bioassays to drug screening. To this end, we investigated the reactions of a panel of 15 luminescent recombinant <I>Escherichia coli</I> bacterial bioreporters to a library of 420 pharmaceuticals. The panel included bacterial bioreporters associated with oxidative stress, DNA damage, heat shock, and efflux of excess metals. Eighty nine drugs elicited a response from at least one of the panel members and formed distinctive clusters, some of which contained closely related drugs. In addition, we tested a group of selected nine drugs against a collection of about 2000 different fluorescent transcriptional reporters that covers the great majority of gene promoters in <I>E. coli.</I> The sets of induced genes were in accord with the in vitro toxicity of the tested drugs, as reflected by the response patterns of the 15-member panel, and provided more insights into their toxicity mechanisms. Facilitated by microplates and robotic systems, all assays were conducted in high-throughput. Our results thus suggest that multi-strain assemblages of bacterial bioreporters have the potential for playing a significant role in drug development alongside current in vitro toxicity tests.</P> <P><B>Highlights</B></P> <P> <UL> <LI> We investigated the reactions of bacterial bioreporters to 420 drug library. </LI> <LI> We tested 9 drugs against transcriptional reporters for majority of gene promoters. </LI> <LI> Results suggest that bioreporters have the potential for drug development. </LI> </UL> </P>
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Elad, Tal,Lee, Jin Hyung,Belkin, Shimshon,Gu, Man Bock Blackwell Publishing Ltd 2008 MICROBIAL BIOTECHNOLOGY -BLACKWELL- Vol.1 No.2
<P><B>Summary</B></P><P>The coming of age of whole‐cell biosensors, combined with the continuing advances in array technologies, has prepared the ground for the next step in the evolution of both disciplines – the whole‐cell array. In the present review, we highlight the state‐of‐the‐art in the different disciplines essential for a functional bacterial array. These include the genetic engineering of the biological components, their immobilization in different polymers, technologies for live cell deposition and patterning on different types of solid surfaces, and cellular viability maintenance. Also reviewed are the types of signals emitted by the reporter cell arrays, some of the transduction methodologies for reading these signals and the mathematical approaches proposed for their analysis. Finally, we review some of the potential applications for bacterial cell arrays, and list the future needs for their maturation: a richer arsenal of high‐performance reporter strains, better methodologies for their incorporation into hardware platforms, design of appropriate detection circuits, the continuing development of dedicated algorithms for multiplex signal analysis and – most importantly – enhanced long‐term maintenance of viability and activity on the fabricated biochips.</P>
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