Dylight 680

Manufactured by Thermo Fisher Scientific

Sourced in United States

DyLight 680 is a fluorescent dye that can be used to label proteins, nucleic acids, and other biological molecules. It has an excitation maximum at 682 nm and an emission maximum at 715 nm, making it suitable for detection in the near-infrared range.

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Lab products found in correlation

Dylight 800, by Thermo Fisher Scientific (11 mentions) Odyssey infrared imaging system, by LI COR (4 mentions) Hmga2, by Cell Signaling Technology (2 mentions) Lin28a, by Cell Signaling Technology (2 mentions) Anti plc γ1 py783, by Cell Signaling Technology (2 mentions) Anti src py416, by Cell Signaling Technology (2 mentions) Anti erk1 2 ptpy185 187, by Thermo Fisher Scientific (2 mentions) Anti gapdh, by Meridian Bioscience (2 mentions) Hexokinase, by Worthington (2 mentions) Dylight 680 nhs ester, by Thermo Fisher Scientific (2 mentions) Blocking solution, by LI COR (2 mentions) Odyssey imager, by LI COR (2 mentions) Odyssey clx, by LI COR (2 mentions) Nanodrop, by Thermo Fisher Scientific (2 mentions) Ripa buffer, by Thermo Fisher Scientific (2 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions) Goat anti rabbit igg h l hrp secondary antibody, by Abcam (1 mentions) M9942, by Merck Group (1 mentions) Sp8 confocal microscope, by Leica (1 mentions) Odyssey clx near infrared fluorescence imaging system, by LI COR (1 mentions)

Close spelling variants of this product

DyLight 680/800 Goat anti-rabbit IgG DyLight 680 conjugate Anti-rabbit IgG–DyLight 680 DyLight™680 conjugated goat anti-rabbit IgG antibody (H&L) Goat anti-mouse IgG DyLight 680 Anti-mouse (IgG) (H+L) Dylight 680 Conjugate Goat anti-mouse DyLight 680 DyLight 680 maleimide DyLight 680 Antibody Labeling Kit DyLight 680 NHS-ester

42 protocols using dylight 680

In Vivo Fluorescence Imaging of Tumor

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The interchain disulfide bond of 7DC2 and 7DC4 were reduced by excess of TCEP at 30°C for 2 hours to produce free sulfhydryl groups. The samples were then conjugated with 10.8 equivalents of DyLight 680 (Thermo Scientific) at 30°C for 4 hours. Unreacted DyLight 680 was removed by using Dye Removal Column kit (Pierce) and conjugation efficiency was determined using Nanodrop (Thermo Scientific) to calculate the molar ratio of DyLight 680 to protein.
Mice bearing 95D tumors were intraperitoneally injected with 0.5 nmol of 7DC2-DyLight 680 or 7DC4-DyLight 680 (100 μL per injection; 3 mg/kg) or 1 nmol of free DyLight680 as a control group. For in vivo optical imaging, NIRF images were obtained at 24 hours using a small-animal IVIS imaging system (IVIS-Spectrum, Xenogen) with excitation and emission wavelengths of 675 and 720 nm. Fluorescence emission was normalized to photons per second per centimeter squared per steradian (p/s/cm2/sr). For ex vivo NIRF imaging, the mice were euthanized 24 hours after injection, and blood and organs were collected. NIRF images were acquired for each tissue as described above.

Chiang Z.C., Fang S., Shen Y.K., Cui D., Weng H., Wang D., Zhao Y., Lin J, & Chen Q. (2022). Development of Novel CD47-Specific ADCs Possessing High Potency Against Non-Small Cell Lung Cancer in vitro and in vivo. Frontiers in Oncology, 12, 857927.

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Fluorescent Secondary Antibody Dilutions

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1:10,000 goat anti-rabbit IgG (H+L) Dylight 800 (ThermoFisher, A32735), 1:10,000 goat anti-mouse IgG (H+L) Dylight 800 (ThermoFisher, A32730), 1:10,000 goat anti-rabbit IgG (H+L) Dylight 680 (ThermoFisher, A32734), 1:10,000 goat anti-mouse IgG (H+L) Dylight 680 (ThermoFisher, A32729), 1:10,000 goat anti-rat IgG (H+L) Dylight 800 (ThermoFisher, sa510024).

Llobet Rosell A., Paglione M., Gilley J., Kocia M., Perillo G., Gasparrini M., Cialabrini L., Raffaelli N., Angeletti C., Orsomando G., Wu P.H., Coleman M.P., Loreto A, & Neukomm L.J. (2022). The NAD+ precursor NMN activates dSarm to trigger axon degeneration in Drosophila. eLife, 11, e80245.

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Quantifying MYOC Protein Levels

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Western blotting of MYOC was performed using the standard protocol and antibodies used in this study were as follows: Anti-MYOC rabbit polyclonal antibody, Sangon Biotech, Order No. D227350; Mouse anti human GAPDH Loading Control Monoclonal Antibody (GA1R), DyLight™ 680, Invitrogen, Product No. MA5-15,738-D680; and Goat anti-Rabbit IgG H&L (HRP) Secondary Antibody, abcam, Cat No. ab6721.

Li H., Han X., Du W., Meng Y., Li Y., Sun T., Liang Q., Li C., Suo C., Gao X., Qiu Y., Tian W., An M., Zhang H., Fu Y., Li X., Lan T., Yang S., Zhang Z., Geng W., Ding C, & Shang H. (2022). Comparative miRNA transcriptomics of macaques and mice reveals MYOC is an inhibitor for Cryptococcus neoformans invasion into the brain. Emerging Microbes & Infections, 11(1), 1572-1585.

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Immunofluorescent Staining of Hippocampal Neurons

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Hippocampal neurons were seeded onto cover slips in the 24-well plates for experimental treatments. Then cells were fixed with 4% paraformaldehyde at room temperature for 15 min, blocked up by PBS supplemented with 5% FBS and 0.3 % Triton X-100 at room temperature for 1 h and finally incubated with primary antibody against MAP2 (dilution 1:500, #M9942, Sigma-Aldrich) at 4 °C overnight. After rinsing in PBS for 3 times to remove free antibodies, cells were incubated with donkey anti-mouse IgG (H + L) cross-adsorbed secondary antibody, dyLight 680 (dilution 1:2000, #SA5−10170, Invitrogen) at room temperature for 1.5 h. Images were captured by Leica SP8 confocal microscope with DAPI (4',6-diamidino-2-phenylindole) for staining of the nuclei.

Yang J.R., Ren T.T., Lan R, & Qin X.Y. (2020). Tea polyphenols attenuate staurosporine-induced cytotoxicity and apoptosis by modulating BDNF-TrkB/Akt and Erk1/2 signaling axis in hippocampal neurons. IBRO Reports, 8, 115-121.

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Western Blot Analysis of Bacterial Proteins

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Bacterial strains were suspended in PBS, then 8 × 10⁶ cells were mixed with SDS-PAGE sample buffer and boiled for 10 min. The samples were then run by electrophoresis on a 10% SDS gel followed by semi-dry transfer onto PVDF membrane. The membranes were blocked with 5% skim milk in tris-buffered saline/Tween (TBST; 50 mM Tris, 150 mM NaCl, 0.1% Tween 20; pH 7.6) for 3 h and incubated with 1 μg/mL of B5 mAb at 4°C overnight. Goat anti-mouse IgG (H+L) cross-absorbed secondary antibody, DyLight 680 (Invitrogen) was used at 1:5,000 dilution to facilitate detection via fluorescent signal. The final development of the signal was detected using the Odyssey CLx Near-Infrared Fluorescence Imaging System (LICOR) and analyzed using the Image Studio software (LICOR).

Wernery U., Chan E., Raghavan R., Teng J.L., Syriac G., Siu S.Y., Joseph M., Yeung M.L., Jia L., Cai J.P., Chiu T.H., Lau S.K, & Woo P.C. (2021). Development of a sensitive competitive enzyme-linked immunosorbent assay for serodiagnosis of Burkholderia mallei, a Tier 1 select agent. PLoS Neglected Tropical Diseases, 15(12), e0010007.

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Characterization of PKR and PACT Interaction

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The pGEX-4T vector expressing the PKR 1–265 fragment in fusion with GST at its N-terminus (GST-PKR Nter) and the pET15b vector expressing the PACT protein have been previously described^{2 (link),15 (link)}. The pENTR™-SD⁄D-TOPO® vector and the pDEST24 were purchased from Invitrogen. Normal mouse or rabbit IgGs were from Santa Cruz Biotechnology. Mouse monoclonal anti-PKR 71/10 antibody was produced by Agrobio Laboratories (FR) as reported^{49 (link)}. Mouse monoclonal Anti-ß-actin antibody was from Sigma (A1978). Rabbit polyclonal antibodies were used to detect human PKR phosphorylated at T446 (ab32036; Abcam). Anti-PACT (sc-377103) and anti-NFκB p65 (sc-109) polyclonal antibodies were purchased from Santa Cruz Biotechnology. Goat anti-Rabbit IgG (H&L) Secondary Antibody, DyLight 800 4X PEG (#SA5-35571), Goat anti-Mouse IgG (H + L) Secondary Antibody, DyLight 680 (#35518) and Alexa Fluor 488 goat anti-rabbit IgG secondary antibody (A11034) were from Invitrogen.

Dabo S., Maillard P., Collados Rodriguez M., Hansen M.D., Mazouz S., Bigot D.J., Tible M., Janvier G., Helynck O., Cassonnet P., Jacob Y., Bellalou J., Gatignol A., Patel R.C., Hugon J., Munier-Lehmann H, & Meurs E.F. (2017). Inhibition of the inflammatory response to stress by targeting interaction between PKR and its cellular activator PACT. Scientific Reports, 7, 16129.

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Immunofluorescence Staining of PDLSCs

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PDLSCs were treated under different conditions on the cell climbing sheets for 1 day. Fixed by paraformaldehyde, 1000 μL blocking buffer (0.2% Triton-100 and 5% donkey serum) was added onto each sheet. Then, the cells were incubated with the primary antibody (VEGF-a or LC3B) at 4°C overnight. After rewarming to room temperature, the cells were dealt with the secondary antibody, DyLight680 (Invitrogen, CA, USA) for 1 hour. DAPI and phalloidin (CoraLite488, Proteintech, Wuhan, China) were used to stain cell nucleus and cytoskeleton. Images of cells were observed and captured under Olympus SpinSR confocal laser microscope (Olympus, Tokyo, Japan).

Huang D., Lei J., Li X., Jiang Z., Luo M, & Xiao Y. (2022). Erythropoietin Activates Autophagy to Regulate Apoptosis and Angiogenesis of Periodontal Ligament Stem Cells via the Akt/ERK1/2/BAD Signaling Pathway under Inflammatory Microenvironment. Stem Cells International, 2022, 9806887.

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Immunoblotting analysis of epithelial-mesenchymal markers

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Cells were lysed, and proteins were separated by SDS-PAGE and transferred to PVDF membrane. After blocking of non-specific binding, immunoblots were incubated with primary antibodies for Snail (L70G2; Cell Signaling Technology, Danvers, MA, USA)^{30 (link)}, E-cadherin (610182; BD Biosciences, San Jose, CA, USA)^{31 (link)}, α/β-tubulin (2148 S; Cell Signaling Technology, Danvers, MA, USA)^{32 (link)}, and GAPDH (14C10; Cell Signaling Technology, Danvers, MA, USA)^{33 (link)} followed by incubation with an anti-mouse IgG conjugated with DyLight 800 (SA5-10176; Invitrogen, Carlsbad, CA, USA)^{34 (link)} or anti-rabbit IgG antibody conjugated with DyLight 680 (35569; Invitrogen, Carlsbad, CA, USA)^{35 (link)}. Immunoblots were scanned and visualized using Odyssey Infrared Imaging System (LI-COR Biosciences, Lincoln, NE, USA). Densitometry was performed on scanned immunoblots by ImageJ.software (National Institutes of Health, Bethesda, MD, USA).

Hojo N., Huisken A.L., Wang H., Chirshev E., Kim N.S., Nguyen S.M., Campos H., Glackin C.A., Ioffe Y.J, & Unternaehrer J.J. (2018). Snail knockdown reverses stemness and inhibits tumour growth in ovarian cancer. Scientific Reports, 8, 8704.

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Immunoblotting Analysis of Protein Expression

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Cells were lysed in Laemmli buffer and sonicated, and proteins were separated by SDS/PAGE and transferred to a PVDF membrane. To prevent nonspecific binding, the membrane was blocked with 0.1%‐5% milk in Tris‐buffered saline with 0.2% Tween (TBST) for 1 h for each immunoblotting antibody. Immunoblotting was performed with primary antibodies including α/β‐tubulin, LIN28A, and HMGA2 (Cell Signaling Technology, Danvers, MA, USA). Secondary antibody immunoblotting was done with anti‐rabbit IgG conjugated with DyLight 680 (Invitrogen, Carlsbad, CA, USA). PVDF membranes were imaged by Odyssey LI‐COR Infrared Imaging System (LI‐COR Biosciences, Lincoln, NE, USA). Densitometry analysis was performed by ImageJ software (National Institutes of Health, Bethesda, MD, USA).

Chirshev E., Hojo N., Bertucci A., Sanderman L., Nguyen A., Wang H., Suzuki T., Brito E., Martinez S.R., Castañón C., Mirshahidi S., Vazquez M.E., Wat P., Oberg K.C., Ioffe Y.J, & Unternaehrer J.J. (2020). Epithelial/mesenchymal heterogeneity of high‐grade serous ovarian carcinoma samples correlates with miRNA let‐7 levels and predicts tumor growth and metastasis. Molecular Oncology, 14(11), 2796-2813.

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Western Blotting of Polyubiquitin Linkages

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To maintain the specificity of ubiquitin linkage-specific antibodies, we followed previously published protocols [25 (link)]. Proteins were resolved by SDS-PAGE in reducing sample buffer, and wet-transferred at 25V for 1 hour to nitrocellulose. Membranes were blocked in 2% non-fat dried milk in TBS-T (0.02% Tween-20) for 1 hour, followed by incubation with primary antibodies (anti-K11 polyubiquitin clone 2A3/2E6 (3 μg/ml), anti-K48 polyubiquitin clone Apu2.07 (1:2000), anti-K63 polyubiquitin clone Apu3 (1:2000)) for 1 hour at room temperature. Membranes were washed three times in TBS-T, followed by incubation with secondary antibodies for 30 minutes at room temperature. Anti-K11 was detected with anti-human IgG IRDye800 (Rockland), and anti-K48/63 with anti-rabbit DyLight 680 (Thermo Scientific), and analyzed on a Licor Odyssey infrared imaging system. Appropriate antigen specificity under these conditions was verified with equimolar amounts of purified diubiquitin molecules (K11, K48, and K63).

Locke M., Toth J.I, & Petroski M.D. (2014). K11- and K48-Linked Ubiquitin Chains Interact with p97 during Endoplasmic Reticulum-Associated Degradation. The Biochemical journal, 459(1), 205-216.

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