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Article ; Online: Engineered EF-Tu and tRNA-Based FRET Screening Assay to Find Inhibitors of Protein Synthesis in Bacteria.

Bhatt, Rachana / Chudaev, Maxim / Mandecki, Wlodek / Goldman, Emanuel

2018 Volume 16, Issue 4, Page(s) 212–221

Abstract: Antibiotic-resistant infections that do not respond to available drugs are becoming more common. Methicillin-resistant Staphylococcus aureus, carbapenem-resistant enterobacteria ("superbugs"), and many others pose a continuous threat to public health. To ...

Abstract	Antibiotic-resistant infections that do not respond to available drugs are becoming more common. Methicillin-resistant Staphylococcus aureus, carbapenem-resistant enterobacteria ("superbugs"), and many others pose a continuous threat to public health. To provide tools to combat such deadly infections, we present in this study a homogeneous assay focused on an insufficiently addressed molecular interaction linked to ribosomal translation. We show that a fluorescence resonance energy transfer (FRET) based screening assay can identify antibiotic molecules that inhibit ternary complex (EF-Tu:tRNA:GTP complex) formation, and therefore, protein synthesis in bacteria. Specifically engineered Escherichia coli EF-Tu and tRNA
MeSH term(s)	Anti-Bacterial Agents/chemistry ; Anti-Bacterial Agents/isolation & purification ; Anti-Bacterial Agents/pharmacology ; Escherichia coli/drug effects ; Escherichia coli/metabolism ; Escherichia coli Proteins/biosynthesis ; Fluorescence Resonance Energy Transfer ; Microbial Sensitivity Tests ; Peptide Elongation Factor Tu/genetics ; Peptide Elongation Factor Tu/isolation & purification ; Peptide Elongation Factor Tu/metabolism ; Protein Biosynthesis/drug effects ; Protein Engineering ; RNA, Transfer/chemistry ; RNA, Transfer/genetics ; RNA, Transfer/isolation & purification
Chemical Substances	Anti-Bacterial Agents ; Escherichia coli Proteins ; RNA, Transfer (9014-25-9) ; Peptide Elongation Factor Tu (EC 3.6.1.-)
Language	English
Publishing date	2018-06-05
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ISSN	1557-8127
ISSN (online)	1557-8127
DOI	10.1089/adt.2018.843
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Article ; Online: Tagging of individual embryos with electronic p-Chips.

Mandecki, Wlodek / Rodriguez, Efrain Frank / Drawbridge, Julie

Biomedical microdevices

2016 Volume 18, Issue 6, Page(s) 100

Abstract: Collecting information about biochemical processes occurring inside a single cell or embryo is traditionally done either using fluorescent dyes with microscopy or via microelectrode voltage-clamp techniques. This paper demonstrates that a more direct ... ...

Abstract	Collecting information about biochemical processes occurring inside a single cell or embryo is traditionally done either using fluorescent dyes with microscopy or via microelectrode voltage-clamp techniques. This paper demonstrates that a more direct method - transmission of information using an electronic chip implanted in an embryo - is feasible. A light-activated microtransponder with dimensions 250 μm × 250 μm × 100 μm (a "p-Chip") was implanted into a blastula-stage frog (Xenopus laevis) embryo. To implant the chip, a small slit is made in the blastocoel roof with an electrolytically-sharpened tungsten needle, and the p-Chip is inserted using fine forceps. The chip is activated when illuminated by a 60 mW focused laser beam, which causes the p-Chip to send its numeric ID to a nearby receiver. At no time during signal transmission does a wire or other type of object come in contact with or penetrate the epidermal layer covering the p-Chip. The embryo survives the procedure, extruding the chip after approximately 3 h. The method shows promise for studies including voltage potential, pH and other parameters.
MeSH term(s)	Animals ; Blastocyst ; Electrical Equipment and Supplies ; Embryo, Nonmammalian/cytology ; Embryo, Nonmammalian/metabolism ; Materials Testing ; Xenopus laevis/embryology
Language	English
Publishing date	2016-12
Publishing country	United States
Document type	Journal Article
ZDB-ID	2004019-2
ISSN	1572-8781 ; 1387-2176
ISSN (online)	1572-8781
ISSN	1387-2176
DOI	10.1007/s10544-016-0127-2
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Article ; Online: FRET study in oligopeptide-linked donor-acceptor system in PVA matrix.

Shah, Sunil / Mandecki, Wlodek / Li, Ji / Gryczynski, Zygmunt / Borejdo, Julian / Gryczynski, Ignacy / Fudala, Rafal

Methods and applications in fluorescence

2016 Volume 4, Issue 4, Page(s) 47002

Abstract: An oligopeptide: Lys-Gly-Pro-Arg-Ser-Leu-Ser-Gly-Lys- ... ...

Abstract	An oligopeptide: Lys-Gly-Pro-Arg-Ser-Leu-Ser-Gly-Lys-NH
MeSH term(s)	Amino Acid Sequence ; Fluorescence Resonance Energy Transfer ; Fluorescent Dyes ; Matrix Metalloproteinase 2 ; Matrix Metalloproteinase 7 ; Matrix Metalloproteinase 9 ; Oligopeptides ; Peptide Fragments ; Peptides
Chemical Substances	Fluorescent Dyes ; Oligopeptides ; Peptide Fragments ; Peptides ; Matrix Metalloproteinase 7 (EC 3.4.24.23) ; Matrix Metalloproteinase 2 (EC 3.4.24.24) ; Matrix Metalloproteinase 9 (EC 3.4.24.35)
Language	English
Publishing date	2016-12-13
Publishing country	England
Document type	Journal Article
ISSN	2050-6120
ISSN (online)	2050-6120
DOI	10.1088/2050-6120/4/4/047002
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Article: Electronic p-Chip-Based System for Identification of Glass Slides and Tissue Cassettes in Histopathology Laboratories.

Mandecki, Wlodek / Qian, Jay / Gedzberg, Katie / Gruda, Maryanne / Rodriguez, Efrain Frank / Nesbitt, Leslie / Riben, Michael

Journal of pathology informatics

2018 Volume 9, Page(s) 9

Abstract: Background: The tagging system is based on a small, electronic, wireless, laser-light-activated microtransponder named "p-Chip." The p-Chip is a silicon integrated circuit, the size of which is 600 μm × 600 μm × 100 μm. Each p-Chip contains a unique ... ...

Abstract	Background: The tagging system is based on a small, electronic, wireless, laser-light-activated microtransponder named "p-Chip." The p-Chip is a silicon integrated circuit, the size of which is 600 μm × 600 μm × 100 μm. Each p-Chip contains a unique identification code stored within its electronic memory that can be retrieved with a custom reader. These features allow the p-Chip to be used as an unobtrusive and scarcely noticeable ID tag on glass slides and tissue cassettes. Methods: The system is comprised of p-Chip-tagged sample carriers, a dedicated benchtop p-Chip ID reader that can accommodate both objects, and an additional reader (the Wand), with an adapter for reading IDs of glass slides stored vertically in drawers. On slides, p-Chips are attached with adhesive to the center of the short edge, and on cassettes - embedded directly into the plastic. ID readout is performed by bringing the reader to the proximity of the chip. Standard histopathology laboratory protocols were used for testing. Results: Very good ID reading efficiency was observed for both glass slides and cassettes. When processed slides are stored in vertical filing drawers, p-Chips remain readable without the need to remove them from the storage location, thereby improving the speed of searches in collections. On the cassettes, the ID continues to be readable through a thin layer of paraffin. Both slides and tissue cassettes can be read with the same reader, reducing the need for redundant equipment. Conclusions: The p-Chip is stable to all chemical challenges commonly used in the histopathology laboratory, tolerates temperature extremes, and remains durable in long-term storage. The technology is compatible with laboratory information management systems software systems. The p-Chip system is very well suited for identification of glass slides and cassettes in the histopathology laboratory.
Language	English
Publishing date	2018-04-02
Publishing country	United States
Document type	Journal Article
ZDB-ID	2579241-6
ISSN	2153-3539 ; 2229-5089
ISSN (online)	2153-3539
ISSN	2229-5089
DOI	10.4103/jpi.jpi_64_17
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Article: Silver nanoparticle-enhanced fluorescence in microtransponder-based immuno- and DNAhybridization assays

Li, Ji / Wang, Zhuying / Gryczynski, Ignacy / Mandecki, Wlodek

Analytical and bioanalytical chemistry. 2010 Nov., v. 398, no. 5

2010

Abstract: The aim of this study is to improve assay sensitivity in common solid-phase bioassay configurations as the result of using silver nanoparticles. The solid phase was provided by numerically indexed, silicon-based electronic chips, microtransponders (p- ... ...

Abstract	The aim of this study is to improve assay sensitivity in common solid-phase bioassay configurations as the result of using silver nanoparticles. The solid phase was provided by numerically indexed, silicon-based electronic chips, microtransponders (p-Chips) that have previously been used in multiplexed assays. Assay configurations investigated included an ELISA-type immunoassay and a DNA hybridization assay. The surface of p-Chips was derivatized with the silver island film (SIF) and a polymer, and then characterized with AFM and SEM. Silver nanoparticle sizes were in the range of 100 to 200 nm. Four fluorophores were tested for fluorescence enhancement; namely, green fluorescent protein, phycoerythrin, Cy3 and Alexa Fluor 555. We consistently observed significant fluorescence enhancement and sensitivity improvement in the p-Chip-based assays: the sensitivity in the cytokine IL-6 immunoassay was 4.3 pg/ml, which represented a 25-fold increase over the method not involving a SIF; and 50 pM in the hybridization assay, a 38-fold increase. The greatest enhancement was obtained for p-Chip surfaces derivatized first with the polymer and then coated with SIF. In conclusion, we show that the SIF-p-Chip-based platform is a highly sensitive method to quantify low-abundance biomolecules in nucleic acid-based assays and immunoassays. [graphic removed]
Keywords	enzyme-linked immunosorbent assay ; hybridization ; surface plasmon resonance
Language	English
Dates of publication	2010-11
Size	p. 1993-2001.
Publisher	Springer-Verlag
Publishing place	Berlin/Heidelberg
Document type	Article
ISSN	1618-2642
DOI	10.1007/s00216-010-4108-7
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: Macrophage inhibitory cytokine 1 biomarker serum immunoassay in combination with PSA is a more specific diagnostic tool for detection of prostate cancer.

Li, Ji / Veltri, Robert W / Yuan, Zhen / Christudass, Christhunesa S / Mandecki, Wlodek

PloS one

2015 Volume 10, Issue 4, Page(s) e0122249

Abstract: Background: Prostate cancer (PCa) is the most common malignancy among men in the United States. Though highly sensitive, the often-used prostate-specific antigen (PSA) test has low specificity which leads to overdiagnosis and overtreatment of PCa. This ... ...

Abstract	Background: Prostate cancer (PCa) is the most common malignancy among men in the United States. Though highly sensitive, the often-used prostate-specific antigen (PSA) test has low specificity which leads to overdiagnosis and overtreatment of PCa. This paper presents results of a retrospective study that indicates that testing for macrophage inhibitory cytokine 1 (MIC-1) concentration along with the PSA assay could provide much improved specificity to the assay. Methods: The MIC-1 serum level was determined by a novel p-Chip-based immunoassay run on 70 retrospective samples. The assay was configured on p-Chips, small integrated circuits (IC) capable of storing in their electronic memories a serial number to identify the molecular probe immobilized on its surface. The distribution of MIC-1 and pre-determined PSA concentrations were displayed in a 2D plot and the predictive power of the dual MIC-1/PSA assay was analyzed. Results: MIC-1 concentration in serum was elevated in PCa patients (1.44 ng/ml) compared to normal and biopsy-negative individuals (0.93 ng/ml and 0.88 ng/ml, respectively). In addition, the MIC-1 level was correlated with the progression of PCa. The area under the receiver operator curve (AUC-ROC) was 0.81 providing an assay sensitivity of 83.3% and specificity of 60.7% by using a cutoff of 0.494 for the logistic regression value of MIC-1 and PSA. Another approach, by defining high-frequency PCa zones in a two-dimensional plot, resulted in assay sensitivity of 78.6% and specificity of 89.3%. Conclusions: The analysis based on correlation of MIC-1 and PSA concentrations in serum with the patient PCa status improved the specificity of PCa diagnosis without compromising the high sensitivity of the PSA test alone and has potential for PCa prognosis for patient therapy strategies.
MeSH term(s)	Aged ; Biomarkers, Tumor/blood ; Biopsy ; Disease Progression ; Growth Differentiation Factor 15/blood ; Humans ; Male ; Middle Aged ; Prognosis ; Prostate-Specific Antigen/blood ; Prostatic Neoplasms/blood ; Prostatic Neoplasms/pathology ; Retrospective Studies
Chemical Substances	Biomarkers, Tumor ; GDF15 protein, human ; Growth Differentiation Factor 15 ; Prostate-Specific Antigen (EC 3.4.21.77)
Language	English
Publishing date	2015
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ISSN	1932-6203
ISSN (online)	1932-6203
DOI	10.1371/journal.pone.0122249
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Article ; Online: Silver nanoparticle-enhanced fluorescence in microtransponder-based immuno- and DNA hybridization assays.

Li, Ji / Wang, Zhuying / Gryczynski, Ignacy / Mandecki, Wlodek

Analytical and bioanalytical chemistry

2010 Volume 398, Issue 5, Page(s) 1993–2001

Abstract	The aim of this study is to improve assay sensitivity in common solid-phase bioassay configurations as the result of using silver nanoparticles. The solid phase was provided by numerically indexed, silicon-based electronic chips, microtransponders (p-Chips) that have previously been used in multiplexed assays. Assay configurations investigated included an ELISA-type immunoassay and a DNA hybridization assay. The surface of p-Chips was derivatized with the silver island film (SIF) and a polymer, and then characterized with AFM and SEM. Silver nanoparticle sizes were in the range of 100 to 200 nm. Four fluorophores were tested for fluorescence enhancement; namely, green fluorescent protein, phycoerythrin, Cy3 and Alexa Fluor 555. We consistently observed significant fluorescence enhancement and sensitivity improvement in the p-Chip-based assays: the sensitivity in the cytokine IL-6 immunoassay was 4.3 pg/ml, which represented a 25-fold increase over the method not involving a SIF; and 50 pM in the hybridization assay, a 38-fold increase. The greatest enhancement was obtained for p-Chip surfaces derivatized first with the polymer and then coated with SIF. In conclusion, we show that the SIF-p-Chip-based platform is a highly sensitive method to quantify low-abundance biomolecules in nucleic acid-based assays and immunoassays.
MeSH term(s)	Electronics/instrumentation ; Fluorescent Dyes ; Immunoassay/methods ; Limit of Detection ; Metal Nanoparticles/chemistry ; Microscopy, Atomic Force ; Silver/chemistry
Chemical Substances	Fluorescent Dyes ; Silver (3M4G523W1G)
Language	English
Publishing date	2010-08-27
Publishing country	Germany
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	201093-8
ISSN	1618-2650 ; 0016-1152 ; 0372-7920
ISSN (online)	1618-2650
ISSN	0016-1152 ; 0372-7920
DOI	10.1007/s00216-010-4108-7
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Article ; Online: Tagging of Test Tubes with Electronic p-Chips for Use in Biorepositories.

Mandecki, Wlodek / Kopacka, Wesley M / Qian, Ziye / Ertwine, Von / Gedzberg, Katie / Gruda, Maryann / Reinhardt, David / Rodriguez, Efrain

Biopreservation and biobanking

2017 Volume 15, Issue 4, Page(s) 293–304

Abstract: A system has been developed to electronically tag and track test tubes used in biorepositories. The system is based on a light-activated microtransponder, also known as a "p-Chip." One of the pressing problems with storing and retrieving biological ... ...

Abstract	A system has been developed to electronically tag and track test tubes used in biorepositories. The system is based on a light-activated microtransponder, also known as a "p-Chip." One of the pressing problems with storing and retrieving biological samples at low temperatures is the difficulty of reliably reading the identification (ID) number that links each storage tube with the database containing sample details. Commonly used barcodes are not always reliable at low temperatures because of poor adhesion of the label to the test tube and problems with reading under conditions of frost and ice accumulation. Traditional radio frequency identification (RFID) tags are not cost effective and are too large for this application. The system described herein consists of the p-Chip, p-Chip-tagged test tubes, two ID readers (for single tubes or for racks of tubes), and software. We also describe a robot that is configured for retrofitting legacy test tubes in biorepositories with p-Chips while maintaining the temperature of the sample below -50°C at all times. The main benefits of the p-Chip over other RFID devices are its small size (600 × 600 × 100 μm) that allows even very small tubes or vials to be tagged, low cost due to the chip's unitary construction, durability, and the ability to read the ID through frost and ice.
MeSH term(s)	Biological Specimen Banks ; Computer Peripherals/economics ; Computer Peripherals/standards ; Radio Frequency Identification Device/economics ; Radio Frequency Identification Device/standards ; Robotics ; Software ; Specimen Handling/instrumentation ; Specimen Handling/standards ; Temperature
Language	English
Publishing date	2017-08
Publishing country	United States
Document type	Journal Article
ZDB-ID	2593993-2
ISSN	1947-5543 ; 1947-5535
ISSN (online)	1947-5543
ISSN	1947-5535
DOI	10.1089/bio.2016.0051
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Article ; Online: Properties of coatings on RFID p-Chips that support plasmonic fluorescence enhancement in bioassays.

Rich, Ryan / Li, Ji / Fudala, Rafal / Gryczynski, Zygmunt / Gryczynski, Ignacy / Mandecki, Wlodek

Analytical and bioanalytical chemistry

2012 Volume 404, Issue 8, Page(s) 2223–2231

Abstract: Microtransponders (RFID p-Chips) derivatized with silver island film (SIF) have previously seen success as a platform for the quantification of low-abundance biomolecules in nucleic acid based assays and immunoassays. In this study, we further ... ...

Abstract	Microtransponders (RFID p-Chips) derivatized with silver island film (SIF) have previously seen success as a platform for the quantification of low-abundance biomolecules in nucleic acid based assays and immunoassays. In this study, we further characterized the morphology of the SIF as well as the polymer matrix enveloping it by scanning electron microscopy (SEM). The polymer was a two-layer silane-based matrix engulfing the p-Chip and SIF. Through a series of SEM and confocal fluorescence microscopy experiments, we found the depth of the polymer matrix to be 1-2 μm. The radiative effects of the SIF/polymer layer were assessed by fluorescence lifetime imaging (FLIM) of p-Chips coated with the polymer to which a fluorophore (Alexa Fluor 555) was conjugated. FLIM images showed an 8.7-fold increase in fluorescence intensity and an increased rate of radiative decay, the latter of which is associated with improved photostability and both of which are linked to plasmonic enhancement by the SIF. Plasmonic enhancement was found to extend uniformly across the p-Chip and, interestingly, to a depth of about 1.2 μm. The substantial depth of enhancement suggests that the SIF/polymer layer constitutes a three-dimensional matrix that is accessible to solvent and small molecules such as fluorescent dyes. Finally, we confirmed that no surface-enhanced Raman scattering is seen from the SIF/polymer combination. The analysis provides a possible mechanism by which the SIF/polymer-coated p-Chips allow a highly sensitive immunoassay and, as a result, leads to an improved bioassay platform.
MeSH term(s)	Biological Assay/instrumentation ; Fluorescence ; Immunoassay ; Microscopy, Electron, Scanning ; Polymers/chemistry ; Radio Frequency Identification Device ; Silver/chemistry ; Surface Plasmon Resonance ; Surface Properties
Chemical Substances	Polymers ; Silver (3M4G523W1G)
Language	English
Publishing date	2012-09-08
Publishing country	Germany
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	201093-8
ISSN	1618-2650 ; 0016-1152 ; 0372-7920
ISSN (online)	1618-2650
ISSN	0016-1152 ; 0372-7920
DOI	10.1007/s00216-012-6369-9
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Article: CCC CGA is a weak translational recoding site in Escherichia coli.

Shu, Ping / Dai, Huacheng / Mandecki, Wlodek / Goldman, Emanuel

Gene

2004 Volume 343, Issue 1, Page(s) 127–132

Abstract: Previously published experiments had indicated unexpected expression of a control vector in which a beta-galactosidase reporter was in the +1 reading frame relative to the translation start. This control vector contained the codon pair CCC CGA in the ... ...

Abstract	Previously published experiments had indicated unexpected expression of a control vector in which a beta-galactosidase reporter was in the +1 reading frame relative to the translation start. This control vector contained the codon pair CCC CGA in the zero reading frame, raising the possibility that ribosomes rephased on this sequence, with peptidyl-tRNA(Pro) pairing with CCC in the +1 frame. This putative rephasing might also be exacerbated by the rare CGA Arg codon in the second position due to increased vacancy of the ribosomal A-site. To test this hypothesis, a series of site-directed mutants was constructed, including mutations in both the first and second codons of this codon pair. The results show that interrupting the continuous run of C residues with synonymous codon changes essentially abolishes the frameshift. Further, changing the rare Arg codon to a common Arg codon also reduces the frequency of the frameshift. These results provide strong support for the hypothesis that CCC CGA in the zero frame is indeed a weak translational frameshift site in Escherichia coli, with a 1-2% efficiency. Because the vector sequence also contains another CCC triplet in the +1 reading frame starting within the next codon after the CGA, our data also support possible contribution to expression of a +7 nucleotide ribosome hop into the same +1 reading frame. We also confirm here a previous report that CCC UGA is a translational frameshift site, in these experiments, with about 5% efficiency.
MeSH term(s)	Amino Acid Sequence ; Base Sequence ; Codon/genetics ; Escherichia coli/genetics ; Frameshift Mutation ; Molecular Sequence Data ; Mutagenesis, Site-Directed ; Protein Biosynthesis ; Restriction Mapping ; Ribosomes/genetics ; beta-Galactosidase/genetics
Chemical Substances	Codon ; beta-Galactosidase (EC 3.2.1.23)
Language	English
Publishing date	2004-12-08
Publishing country	Netherlands
Document type	Journal Article ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	391792-7
ISSN	1879-0038 ; 0378-1119
ISSN (online)	1879-0038
ISSN	0378-1119
DOI	10.1016/j.gene.2004.08.012
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