Cyclooctatetraene

Manufactured by Merck Group

Cyclooctatetraene is a cyclic hydrocarbon compound with the chemical formula C₈H₈. It is a colorless, volatile liquid that is used as a laboratory reagent and in the synthesis of other organic compounds.

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Lab products found in correlation

Phosphocreatine, by Merck Group (2 mentions) 4 nitrobenzyl alcohol, by Merck Group (2 mentions) Protocatechuate 3 4 dioxygenase, by Merck Group (2 mentions) Streptavidin, by Thermo Fisher Scientific (2 mentions) Trolox, by Merck Group (2 mentions) Protocatechuic acid, by Merck Group (2 mentions) Glucose oxidase, by Merck Group (2 mentions) Mercaptoethanol, by Merck Group (1 mentions) Pyranose oxidase, by Merck Group (1 mentions) Fluoroshield, by Thermo Fisher Scientific (1 mentions) Zen software, by Zeiss (1 mentions) Cysteamine, by Merck Group (1 mentions) Creatine kinase, by Merck Group (1 mentions) Digitonin, by Merck Group (1 mentions) Ix83 celltirf, by Olympus (1 mentions) Biotin peg, by Laysan Bio (1 mentions) Flash 4.0 v3, by Hamamatsu Photonics (1 mentions) Metamorph software, by Molecular Devices (1 mentions) Creatine phosphokinase, by Merck Group (1 mentions) Biotinylated bovine serum albumin, by Merck Group (1 mentions)

6 protocols using cyclooctatetraene

Super-Resolution Imaging Protocols: SIM and PALM

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For structured illumination microscopy (SIM) image acquisition, labeled cells on glass coverslips were mounted with Fluoroshield™, containing DAPI (Thermo Fisher). Images were recorded with the 40 × alpha 1.46 Plan apochromat objective with oil immersion (Zeiss, Oberkochen, Germany). Z-stacks were recorded in SIM mode with a 16 bit depth at 5 angles, with averaging 4; 51 µm grid was applied for 633 laser line. The acquired SIM dataset were reconstructed by the ZEN software (Zeiss).
To perform PALM imaging, labeled cells were kept in an imaging solution containing 10% Glucose, 10 mM sodium chloride, 50 mM Tris–HCl, catalase, Pyranose oxidase, 100 mM Cysteamine, 2 mM Cyclooctatetraene and 100 mM Mercaptoethanol (all Sigma Aldrich). The detailed process of image acquisition was reported previously [21 (link)]. A total of 5000–10,000 frames were acquired using a Zeiss ELYRA LSM 780 imaging system (Zeiss) equipped with a 1.57 N.A. 100 × oil objective. Subsequently, reconstruction of raw images and post-processing was performed with Image J software and the Thunderstorm plugin [22 (link)].

Skorska A., Johann L., Chabanovska O., Vasudevan P., Kussauer S., Hillemanns M., Wolfien M., Jonitz-Heincke A., Wolkenhauer O., Bader R., Lang H., David R, & Lemcke H. (2022). Monitoring the maturation of the sarcomere network: a super-resolution microscopy-based approach. Cellular and Molecular Life Sciences, 79(3), 149.

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Single-Molecule Imaging of Multidrug Resistance Protein

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Single-molecule experiments were performed at room temperature (23 ± 1°C) on an objective-type total-internal-reflection fluorescence microscope (Olympus IX83 cellTIRF). Microfluidic imaging chambers were passivated with a mixture of PEG and biotin-PEG (Laysan Bio), and incubated with 0.8 µM streptavidin (Invitrogen) followed by 2 nM fluorescently labeled, His-tagged bMRP1 that had been preincubated with biotinylated anti-His₆ antibodies (Invitrogen) for 1 hr on ice in a buffer containing 50 mM Tris-HCl pH 8.0, 150 mM KCl, 2 mM MgCl₂, 0.06% digitonin, 0.5 mg/mL BSA, 10 mM phosphocreatine (Sigma), and 0.1 mg/mL creatine kinase (Sigma). A triplet-state quenching cocktail (Dave et al., 2009 (link)) of 1 mM cyclooctatetraene (Sigma), 1 mM 4-nitrobenzyl alcohol (Sigma), and 1 mM Trolox (Sigma), as well as an oxygen scavenging system (Aitken et al., 2008 (link)) containing 10 nM protocatechuate-3,4-dioxygenase (Sigma) and 2.5 mM protocatechuic acid (Sigma) were supplemented to the imaging buffer. ATP and/or LTC₄ were included in the imaging buffer at concentrations specified in the text. Fluorescence signals were split with a W-View Gemini-2C (Hamamatsu), directed to two CMOS cameras (Flash 4.0 v3, Hamamatsu), and acquired by MetaMorph software (Molecular Devices) at a frame rate specified in the text.

Wang L., Johnson Z.L., Wasserman M.R., Levring J., Chen J, & Liu S. (2020). Characterization of the kinetic cycle of an ABC transporter by single-molecule and cryo-EM analyses. eLife, 9, e56451.

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Visualizing DNA-ORC-Cdc6 Interactions

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Optical traps tethering a single bare DNA or nucleosomal DNA were moved to a separate channel containing 2 nM Cy3- or LD650-ORC and 5 nM Cdc6 in an imaging buffer containing 25 mM Tris-OAc pH 7.5, 5% glycerol, 40 µg/ml BSA, 3 mM DTT, 2 mM TCEP, 0.1 mM EDTA, 10 mM Mg(OAc)₂, 50 mM potassium glutamate, and 2.5 mM ATP. The imaging buffer was supplemented with an ATP-regeneration system [60 mg/ml creatine phosphokinase (Sigma) and 20 mM phosphocreatine (Sigma)], a triplet-state quenching cocktail [1 mM cyclooctatetraene (Sigma), 1 mM 4-nitrobenzyl alcohol (Sigma) and 1 mM Trolox (Sigma)], as well as an oxygen scavenging system [10 nM protocatechuate-3,4-dioxygenase (Sigma) and 2.5 mM protocatechuic acid (Sigma)]. Kymographs were typically recorded for 4–10 min.

Li S., Wasserman M.R., Yurieva O., Bai L., O’Donnell M.E, & Liu S. (2022). Nucleosome-directed replication origin licensing independent of a consensus DNA sequence. Nature Communications, 13, 4947.

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Quartz Slide Preparation for Single-Molecule Studies

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Home-built quartz slides were prepared as previously described^{36 (link)} with 5 minute epoxy (Staples) used for sealing the edges of the slide. The surface of the slides was pre-functionalized by brief incubation with biotinylated bovine serum albumin (Sigma, 1 mg/ml, 5 minutes), washing, and followed by incubating with streptavidin (Invitrogen, 0.1 mg/ml, 20 minutes) and washing. For slides utilizing heparin attachment, biotin-heparin (Sigma) was either added directly to the slides or pre-incubated with the LPL before addition. Immediately preceding data collection, LPL was deposited onto the slide surface in Buffer A (20 mM Tris, 300 mM sodium chloride, 5% glycerol, 1 mM deoxycholate, pH 7.5) and incubated for 5 minutes. Unbound LPL was then washed out with imaging buffer (Buffer A with the addition of ~50 μM cyclooctatetraene (Sigma), a triplet state quencher to lessen fluorophore blinking.)

Hayne C.K., Yumerefendi H., Cao L., Gauer J.W., Lafferty M.J., Kuhlman B., Erie D.A, & Neher S.B. (2018). We FRET So You Don’t Have To: New Models of the Lipoprotein Lipase Dimer. Biochemistry, 57(2), 241-254.

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STORM Imaging Buffer Optimization

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STORM imaging buffer for cultured cells contains 10% (w/v) glucose (Sigma-Aldrich), glucose oxidase (0.56 mg/ml; Sigma-Aldrich), catalase (0.17 mg/ml; Sigma-Aldrich), and 0.14 M 2-mercaptoethanol (βME, Sigma-Aldrich). For the formalin-fixed paraffin-embedded (FFPE) tissue section, to reduce the high background caused by the strong scattering of pathological tissue, an optical clearing process was conducted before imaging by immersing the sample in 60% (v/v) 2,2′-thiodiethanol (TDE) for 20 to 30 min to make the sample transparent (27 (link)). For the STORM imaging buffer of FFPE tissue section, 60% (v/v) TDE solution was used instead of water to match the tissue’s index and contains 10% (w/v) glucose (Sigma-Aldrich), glucose oxidase (0.56 mg/ml; Sigma-Aldrich), catalase (0.17 mg/ml; Sigma-Aldrich), 0.14 M βME (Sigma-Aldrich), and 0.2 mM cyclooctatetraene (Sigma-Aldrich). The imaging buffer was added to the sample dish right before imaging.

Ma H., Chen M., Nguyen P, & Liu Y. (2024). Toward drift-free high-throughput nanoscopy through adaptive intersection maximization. Science Advances, 10(21), eadm7765.

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Super-Resolution Imaging of Sialic Acids

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dSTORM imaging was performed on Nikon inverted Eclipse Ti-E with a 100 × 1.49 N.A TIRF lens (Nikon, Tokyo, Japan), an objective TIRF illumination. The samples were imaged in freshly prepared buffer containing 10% glucose (Sigma, St. Louis, United States), 50 mM Tris-HCL pH8.0 (Sigma, St. Louis, United States), 10 mM NaCl (Sigma, St. Louis, United States), 2 mM cyclooctatetraene (Sigma, St. Louis, United States), 143 mM beta-mercaptoethanol (Sigma, St. Louis, United States), 0.65 mg/mL glucose oxidase (Sigma, St. Louis, United States) and 0.04 mg/mL catalase from Aspergillus niger (Sigma, St. Louis, United States). The coverslip that containing the seeded cells was sealed onto a microscopic depression slide (Sail Brand, China) using nail polish. The images were captured with excitation of a 640 nm laser with using an excitation filter (ZET532/647x, Chroma), a dichroic mirror (T760LPXR-UF2, Chroma) and an emission filter set (FF01-692/40–25, 25 mm, Semrock, NY, United States). Camera model, iXon Ultra 897 EMCCD (Andor Technology, Belfast, UK). A time series of 10,000 or 20,000 frames per cell was recorded for α-2,3-linked sialic acid and α-2,6-linked sialic acid. During the acquisition time, focus lock for x-y drift was applied with polystyrene beads pre-coated on the coverslip.

Fong C.H., Lu L., Chen L.L., Yeung M.L., Zhang A.J., Zhao H., Yuen K.Y, & To K.K. (2022). Interferon-gamma inhibits influenza A virus cellular attachment by reducing sialic acid cluster size. iScience, 25(4), 104037.

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