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Single molecule imaging

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https://www.readbyqxmd.com/read/29328231/aberration-accounting-calibration-for-3d-single-molecule-localization-microscopy
#1
Clément Cabriel, Nicolas Bourg, Guillaume Dupuis, Sandrine Lévêque-Fort
We propose a straightforward sample-based technique to calibrate the axial detection in 3D single-molecule localization microscopy. Using microspheres coated with fluorescent molecules, the calibration curves of point spread function-shaping or intensity-based measurements can be obtained over the imaging depth range. This experimental method takes into account the effect of the spherical aberration without requiring computational correction. We demonstrate its efficiency for astigmatic imaging in a 1.2 μm range above the coverslip...
January 15, 2018: Optics Letters
https://www.readbyqxmd.com/read/29327523/association-of-physicians-of-india-position-statement-on-role-of-chirally-pure-molecules-in-clinical-practice
#2
Milind Y Nadkar, Mangesh Tiwaskar, Sanjay Kalra, Siddharth N Shah, B R Bansode, Anjanlal Dutta, Sarita Bajaj, Sameer Aggarwal, Yatan Pal Singh Balhara, A K Das, Puneet Dhamija, Y K Gupta, Jubbin Jacob, Sundeep Mishra, S N Narasingan, C K Ponde, Ram Prabhoo, Balakrishnan S, Manisha Sahay, R K Sahay, I Sathyamurthy, Shilpa Tiwaskar, Agam Vora
Chirally pure molecules or enantiomers are non-superimposable mirror images of each other with a chiral center (such as carbon, sulphur, nitrogen or phosphorous atom). An equimolar mixture of enantiomers forms a racemate. Chirally pure molecules (single enantiomers) are important in the field of drug discovery as the drug targets such as enzymes and receptors are enantioselective in nature. Clinical studies have demonstrated that chirally pure drugs exhibit different pharmacokinetic and metabolic profiles, reduced adverse events, improved safety profiles and similar therapeutic activity at lowered drug dosage as compared with the racemate in many therapeutic areas...
December 2017: Journal of the Association of Physicians of India
https://www.readbyqxmd.com/read/29322442/single-molecule-fluorescence-in-situ-hybridization-fish-of-circular-rna-cdr1as
#3
Christine Kocks, Anastasiya Boltengagen, Monika Piwecka, Agnieszka Rybak-Wolf, Nikolaus Rajewsky
Individual mRNA molecules can be imaged in fixed cells by hybridization with multiple, singly labeled oligonucleotide probes, followed by computational identification of fluorescent signals. This approach, called single-molecule RNA fluorescence in situ hybridization (smRNA FISH), allows subcellular localization and absolute quantification of RNA molecules in individual cells. Here, we describe a simple smRNA FISH protocol for two-color imaging of a circular RNA, CDR1as, simultaneously with an unrelated messenger RNA...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29317629/3d-single-molecule-super-resolution-microscopy-with-a-tilted-light-sheet
#4
Anna-Karin Gustavsson, Petar N Petrov, Maurice Y Lee, Yoav Shechtman, W E Moerner
Tilted light sheet microscopy with 3D point spread functions (TILT3D) combines a novel, tilted light sheet illumination strategy with long axial range point spread functions (PSFs) for low-background, 3D super-localization of single molecules as well as 3D super-resolution imaging in thick cells. Because the axial positions of the single emitters are encoded in the shape of each single-molecule image rather than in the position or thickness of the light sheet, the light sheet need not be extremely thin. TILT3D is built upon a standard inverted microscope and has minimal custom parts...
January 9, 2018: Nature Communications
https://www.readbyqxmd.com/read/29317342/imaging-guided-photo-therapeutic-nanoporphyrin-synergized-hsp90-inhibitor-in-patient-derived-xenograft-bladder-cancer-model
#5
Qilai Long, Tzu-Yin Lin, Yee Huang, Xiaocen Li, Ai-Hong Ma, Hongyong Zhang, Randy Carney, Susan Airhart, Kit S Lam, Chong-Xian Pan, Yuanpei Li
Photodynamic therapy is a promising and effective non-invasive therapeutic approach for the treatment of bladder cancers. Therapies targeting HSP90 have the advantage of tumor cell selectivity and have shown great preclinical efficacy. In this study, we evaluated a novel multifunctional nanoporphyrin platform loaded with an HSP90 inhibitor 17AAG (NP-AAG) for use as a multi-modality therapy against bladder cancer. NP-AAG was efficiently accumulated and retained at bladder cancer patient-derived xenograft (PDX) over 7 days...
January 6, 2018: Nanomedicine: Nanotechnology, Biology, and Medicine
https://www.readbyqxmd.com/read/29314718/new-frontiers-in-rna-transport-and-local-translation-in-neurons
#6
REVIEW
S J Van Driesche, K C Martin
RNA localization to neuronal dendrites and axons is increasingly recognized as a significant and widespread mechanism of gene expression control in neurons. High-throughput RNA sequencing is rapidly expanding the universe of known localized mRNAs. Although there are inherent difficulties in preparing sequencing libraries from dendrites and axons in the context of intact brain, genetic labeling strategies have paved the way for improved studies of this type. As the list of localized mRNAs grows, there is increasing need for functional validation of localized transcripts - that is, do particular localized transcripts serve demonstrable physiologic functions in axons or dendrites? Finally, specific details about what localized mRNAs do once they reach distal processes have long been elusive...
January 5, 2018: Developmental Neurobiology
https://www.readbyqxmd.com/read/29308904/pet-imaging-of-vcam-1-expression-and-monitoring-therapy-response-in-tumor-with-a-68ga-labeled-single-chain-variable-fragment
#7
Xiao Zhang, Chunbao Liu, Fan Hu, Yingying Zhang, Jing Wang, Yongheng Gao, Yaqun Jiang, Yongxue Zhang, Xiaoli Lan
Vascular cell adhesion molecule-1 (VCAM-1) is becoming an attractive candidate for targeting in tumor detection and therapy due to its involvement in tumorigenicity and metastasis. LY2409881, an IKK2 inhibitor, can trigger apoptosis of VCAM-1 positive melanoma cells. The aim of this study was to assess the use of the 68Ga-labeled single chain variable fragment (scFv) of VCAM-1 in detecting VCAM-1-positive tumor expression and monitoring the therapeutic effect of LY2409881. Melanoma cell lines, B16F10 and A375m, were sued as VCAM-1 overexpression and underexpression models, respectively...
January 8, 2018: Molecular Pharmaceutics
https://www.readbyqxmd.com/read/29307468/high-free-energy-barrier-of-1d-diffusion-along-dna-by-architectural-dna-binding-proteins
#8
Kiyoto Kamagata, Mano Eriko, Kana Ouchi, Saori Kanbayashi, Reid C Johnson
Architectural DNA-binding proteins function to regulate diverse DNA reactions and have the defining property of significantly changing DNA conformation. Although the 1D movement along DNA by other types of DNA-binding proteins has been visualized, the mobility of architectural DNA-binding proteins on DNA remains unknown. Here, we applied single-molecule fluorescence imaging on arrays of extended DNA molecules to probe the binding dynamics of three structurally-distinct architectural DNA-binding proteins: Nhp6A, HU, and Fis...
January 4, 2018: Journal of Molecular Biology
https://www.readbyqxmd.com/read/29307175/an-ultrasensitive-fret-sef-probe-for-sensing-and-imaging-micrornas-in-living-cells-based-on-gold-nano-conjugates
#9
Jiadi Sun, Fuwei Pi, Jian Ji, Hongtao Lei, Zhixian Gao, Yinzhi Zhang, Jean de Dieu Habimana, Zaijun Li, Xiulan Sun
MicroRNAs (miRNAs), a kind of single-stranded small RNA molecules, play significant roles in the physiological and pathological processes of human beings. Currently, miRNAs had been demonstrated as important biomarkers critically related to many diseases and life nature,including several cancers and cell senescence. It is valuable to establish sensitive assays for monitoring the levels of intracellular up-regulated/down-regulated miRNA expression, which would contribute to the early prediction of the tumor risk and cardiovascular disease...
January 8, 2018: Analytical Chemistry
https://www.readbyqxmd.com/read/29306438/revealing-the-raft-domain-organization-in-the-plasma-membrane-by-single-molecule-imaging-of-fluorescent-ganglioside-analogs
#10
Kenichi G N Suzuki, Hiromune Ando, Naoko Komura, Miku Konishi, Akihiro Imamura, Hideharu Ishida, Makoto Kiso, Takahiro K Fujiwara, Akihiro Kusumi
Gangliosides have been implicated in a variety of physiological processes, particularly in the formation and function of raft domains in the plasma membrane. However, the scarcity of suitable fluorescent ganglioside analogs had long prevented us from determining exactly how gangliosides perform their functions in the live-cell plasma membrane. With the development of new fluorescent ganglioside analogs, as described by Komura et al. (2017), this barrier has been broken. We can now address the dynamic behaviors of gangliosides in the live-cell plasma membrane, using fluorescence microscopy, particularly by single-fluorescent molecule imaging and tracking...
2018: Methods in Enzymology
https://www.readbyqxmd.com/read/29303582/spectroscopic-study-of-a-photoactive-antibacterial-agent-2-3-distyrylindole
#11
Ruwini Dinushika Rajapaksha, Matthias Banet, Cody Champion, Liliya V Frolova, Snezna Rogelj, Pabitra Choudhury, Mahinda Iroshana Ranasinghe
Optical properties and fluorescence decay dynamics of a photoactive indole based antibacterial chromophore system, 2, 3-distyrylindole (23DSI), was investigated using various spectroscopic characterization techniques. These studies were done by utilizing the steady state UV-Vis spectroscopy, the steady state fluorescence spectroscopy, the time-resolve fluorescence upconversion spectroscopy, and time correlated single photon counting spectroscopy as experiment techniques. Our studies show that the 23DSI molecule has a multiphoton absorbing property as indicated by two and three photon absorption in both solution and solid phases...
January 5, 2018: Journal of Physical Chemistry. A
https://www.readbyqxmd.com/read/29303250/we-fret-so-you-don-t-have-to-new-models-of-the-lipoprotein-lipase-dimer
#12
Cassandra K Hayne, Hayretin Yumerefendi, Lin Cao, Jacob W Gauer, Michael J Lafferty, Brian Kuhlman, Dorothy A Erie, Saskia B Neher
Lipoprotein lipase (LPL) is a dimeric enzyme that is responsible for clearing triglyceride-rich lipoproteins from the blood. Although LPL plays a key role in cardiovascular health, an experimentally derived three-dimensional structure has not been determined. Such a structure would aid in understanding mutations in LPL that cause familial LPL deficiency in patients and help in the development of therapeutic strategies to target LPL. A major obstacle to structural studies of LPL is that LPL is an unstable protein that is difficult to produce in the quantities needed for nuclear magnetic resonance or crystallography...
January 5, 2018: Biochemistry
https://www.readbyqxmd.com/read/29300081/branched-hybridization-chain-reaction-bhcr-circuit-for-ultrasensitive-localizable-imaging-of-mrna-in-living-cells
#13
Lan Liu, Jin-Wen Liu, Han Wu, Xiang-Nan Wang, Ru-Qin Yu, Jian-Hui Jiang
Hybridization chain reaction (HCR) circuits are valuable approaches to monitor low-abundance mRNA, and current HCR is still subjected to issues such as limited amplification efficiency, compromised localization resolution or complicated designs. We report a novel branched HCR (bHCR) circuit for efficient signal-amplified imaging of mRNA in living cells. The bHCR can be realized using a simplified design by hierarchically coupling two HCR circuits with two split initiator fragments of the secondary HCR circuit incorporated in the probes for the primary HCR circuit...
January 4, 2018: Analytical Chemistry
https://www.readbyqxmd.com/read/29291166/two-color-based-nanoflares-for-multiplexed-micrornas-imaging-in-live-cells
#14
Jing Li, Jin Huang, Xiaohai Yang, Yanjing Yang, Ke Quan, Nuli Xie, Yanan Wu, Changbei Ma, Kemin Wang
MicroRNAs (miRNAs) have become an ideal biomarker candidate for early diagnosis of diseases. But various diseases involve changes in the expression of different miRNAs. Therefore, multiplexed assay of miRNAs in live cells can provide critical information for our better understanding of their roles in cells and further validating of their function in clinical diagnoses. Simultaneous detection of multiple biomarkers could effectively improve the accuracy of early cancer diagnosis. Here, we develop the two-color-based nanoflares for simultaneously detecting two distinct miRNA targets inside live cells...
2018: Nanotheranostics
https://www.readbyqxmd.com/read/29290471/resolving-the-gordian-knot-srs2-strips-intermediates-formed-during-homologous-recombination
#15
Harshad Ghodke, Jacob S Lewis, Antoine M van Oijen
Cells use a suite of specialized enzymes to repair chromosomal double-strand breaks (DSBs). Two recent studies describe how single-molecule fluorescence imaging techniques are used in the direct visualization of some of the key molecular steps involved. De Tullio et al. and Kaniecki et al. watch individual Srs2 helicase molecules disrupt repair intermediates formed by RPA, Rad51, and Rad52 on DNA during homologous recombination.
December 28, 2017: Trends in Biochemical Sciences
https://www.readbyqxmd.com/read/29287345/mass-spectrometric-imaging-of-localization-of-fat-molecules-in-water-in-oil-emulsions-containing-semi-solid-fat
#16
Makoto Shiota, Yuichi Shimomura, Mariko Kotera, Shu Taira
Firstly, we report the localization analysis of the lipid components of a water-in-oil (W/O) semi-solid emulsion by mass spectrometry imaging (MSI). Uniform emulsion droplets were prepared using microchannel emulsification devices with lecithin, stearic acid-binding monoglyceride (St-MAG), and polyglycerol polyricinoleate (PGPR) as emulsifiers. The mass image gives us the localization of phosphatidylcholine (PC) in lecithin, St-MAG, tripalmitin (PPP), medium-chain triglyceride (MCT), and high-melting-point triglyceride tristearin (C18-TAG)...
April 15, 2018: Food Chemistry
https://www.readbyqxmd.com/read/29286574/internal-rulers-to-assess-fluorescent-protein-photoactivation-efficiency
#17
Malte Renz, Christian Wunder
Photoactivatable fluorescent proteins (PA-FPs) have been widely used to assess the dynamics of cell biological processes. In addition, PA-FPs enabled single-molecule based super-resolution imaging (photoactivated localization microscopy) and thereby provided unprecedented structural insight. For the lack of tools, however, the fraction of PA-FPs that is, actually being switched on to fluoresce, that is, the photoactivation efficiency, has been difficult to assess. Uncertainty about photoactivation efficiency has hampered an understanding of the absolute amount of PA-FPs, that is, being examined...
December 29, 2017: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
https://www.readbyqxmd.com/read/29286479/method-for-labeling-transcripts-in-individual-escherichia-coli-cells-for-single-molecule-fluorescence-in-situ-hybridization-experiments
#18
Rinat Arbel-Goren, Yonatan Shapira, Joel Stavans
A method is described for labeling individual messenger RNA (mRNA) transcripts in fixed bacteria for use in single-molecule fluorescence in situ hybridization (smFISH) experiments in E. coli. smFISH allows the measurement of cell-to-cell variability in mRNA copy number of genes of interest, as well as the subcellular location of the transcripts. The main steps involved are fixation of the bacterial cell culture, permeabilization of cell membranes, and hybridization of the target transcripts with sets of commercially available short fluorescently-labeled oligonucleotide probes...
December 21, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/29284750/cyclic-amp-dependent-plasticity-underlies-rapid-changes-in-odor-coding-associated-with-reward-learning
#19
Thierry Louis, Aaron Stahl, Tamara Boto, Seth M Tomchik
Learning and memory rely on dopamine and downstream cAMP-dependent plasticity across diverse organisms. Despite the central role of cAMP signaling, it is not known how cAMP-dependent plasticity drives coherent changes in neuronal physiology that encode the memory trace, or engram. In Drosophila, the mushroom body (MB) is critically involved in olfactory classical conditioning, and cAMP signaling molecules are necessary and sufficient for normal memory in intrinsic MB neurons. To evaluate the role of cAMP-dependent plasticity in learning, we examined how cAMP manipulations and olfactory classical conditioning modulate olfactory responses in the MB with in vivo imaging...
December 28, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/29284749/visualizing-long-term-single-molecule-dynamics-in-vivo-by-stochastic-protein-labeling
#20
Hui Liu, Peng Dong, Maria S Ioannou, Li Li, Jamien Shea, H Amalia Pasolli, Jonathan B Grimm, Patricia K Rivlin, Luke D Lavis, Minoru Koyama, Zhe Liu
Our ability to unambiguously image and track individual molecules in live cells is limited by packing of multiple copies of labeled molecules within the resolution limit. Here we devise a universal genetic strategy to precisely control copy number of fluorescently labeled molecules in a cell. This system has a dynamic range of ∼10,000-fold, enabling sparse labeling of proteins expressed at different abundance levels. Combined with photostable labels, this system extends the duration of automated single-molecule tracking by two orders of magnitude...
December 28, 2017: Proceedings of the National Academy of Sciences of the United States of America
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