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Irdye 680rd

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IRDye 680RD is a near-infrared fluorescent dye. It has an absorption maximum at 676 nm and an emission maximum at 694 nm. This dye can be used for labeling proteins, nucleic acids, and other biomolecules for detection and analysis in various applications.

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Lab products found in correlation

Irdye 800cw, by LI COR (213 mentions) Odyssey blocking buffer, by LI COR (32 mentions) Odyssey infrared imaging system, by LI COR (30 mentions) Odyssey clx imaging system, by LI COR (28 mentions) Odyssey imaging system, by LI COR (27 mentions) Ripa buffer, by Thermo Fisher Scientific (18 mentions) Pierce bca protein assay kit, by Thermo Fisher Scientific (17 mentions) Nitrocellulose membrane, by Bio-Rad (15 mentions) Odyssey clx, by LI COR (14 mentions) Odyssey clx imager, by LI COR (12 mentions) Odyssey clx system, by LI COR (9 mentions) Immobilon fl, by Merck Group (9 mentions) Nupage 4 12 bis tris gel, by Thermo Fisher Scientific (9 mentions) Alexa fluor 488, by Thermo Fisher Scientific (9 mentions) Immobilon fl pvdf membrane, by Merck Group (9 mentions) Irdye 800rd, by LI COR (8 mentions) Odyssey scanner, by LI COR (8 mentions) Image studio software, by LI COR (8 mentions) Pvdf membrane, by Merck Group (8 mentions) β actin, by Merck Group (7 mentions)

Spelling variants of this product

IRDye 680RD goat anti-rabbit (108 citations) Anti-rabbit IRDye 680RD (17 citations) IRDye 680RD Streptavidin (11 citations) IRDye 680RD-conjugated goat anti-mouse IgG (5 citations) IRDye 680RD goat anti-mouse antibodies (4 citations) Goat α-mouse IRDye 680RD (4 citations) IRDye 680RD donkey anti-mouse IgG antibody (2 citations) IRDye 680RD conjugated second antibodies (2 citations) IRDye 680RD-conjugated goat anti-mouse (2 citations) Donkey anti-mouse IRDye 800CW and goat anti-rabbit IRDye 680RD secondary antibodies (2 citations)

357 protocols using irdye 680rd

1

Immunoblotting: Antibodies for Protein Detection

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Monoclonal antibodies against GFP (clone B-2, 1:2,000 dilution) and Ubiquitin (clone P4D1, 1:2,000 dilution) were purchased from Santa Cruz Biotechnology, monoclonal Pgk1 antibodies (clone 22C5D8, 1:5,000 dilution) were from Invitrogen and monoclonal Ydj1 antibodies (1:5,000 dilution) were from Sigma Aldrich. Monoclonal antibodies against GFP (Clone B34, 1:2,000 dilution) were used to detect the n-terminal part of split-venus (VN). Polyclonal Hsp104 antibodies (1:1,000 dilution) were purchased from Enzo Life sciences and polyclonal Sis1 antibodies (1:10,000 dilution) were from Cosmo Bio. Polyclonal Apj1 antibodies were raised in rabbit against full length Apj1. Fluorescently labeled secondary antibodies (IRDye 800CW, anti-mouse IgG (goat) and IRDye 800CW, anti-rabbit (goat), IRDye 680RD, anti-mouse IgG (goat) and IRDye 680RD, anti-rabbit (goat); each used at 1:10,000 for immunodetection using a Li-Cor system) were from Li-Cor.
den Brave F., Cairo L.V., Jagadeesan C., Ruger-Herreros C., Mogk A., Bukau B, & Jentsch S. (2020). Chaperone-Mediated Protein Disaggregation Triggers Proteolytic Clearance of Intra-nuclear Protein Inclusions. Cell Reports, 31(9), 107680.
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2

Hippocampal Protein Analysis in EHT-Treated Mice

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After 4 weeks of dietary EHT administration animals were euthanized by cervical dislocation–an AVMA approved method that allows for rapid removal of non-hypoxic brains. Hippocampi were then rapidly dissected, snap frozen and stored prior to homogenization for western blot analysis. Hippocampal homogenates were prepared by sonication at 95°C in aqueous buffer containing 2% lithium dodecyl sulfate and 50 mM Tris pH 7.5. Total protein concentrations were determined by bicinchoninic acid assay according to the manufacturer’s instructions (Pierce) and 15 μg of total protein was loaded per lane on SDS-PAGE gels. Proteins were transferred to PVDF membranes, which were then blocked with Odyssey Blocking Buff (TBS) (LI-COR) for 1 hr at rm temp. Blots were probed with primary antibodies (Table 1) overnight followed by the corresponding secondary, which was one of the follow: Goat anti-rabbit (IRDye 800CW LI-COR), Goat anti-mouse (IRDye 680RD LI-COR), or Donkey anti-goat (IRDye 680RD LI-COR). Protein bands were detected by an Odyssey Clx and quantified by ImageStudio. Band intensities were determined for each antigen and normalized to the corresponding within-lane loading control. Data were presented as the mean normalized band intensity for each treatment group ±SEM and expressed as the percent of the mean value for the vehicle-treated control group.
Asam K., Staniszewski A., Zhang H., Melideo S.L., Mazzeo A., Voronkov M., Huber K.L., Pérez E., Stock M., Stock J.B., Arancio O, & Nicholls R.E. (2017). Eicosanoyl-5-hydroxytryptamide (EHT) prevents Alzheimer’s disease-related cognitive and electrophysiological impairments in mice exposed to elevated concentrations of oligomeric beta-amyloid. PLoS ONE, 12(12), e0189413.
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3

Mouse ADRB2 Signaling Assay Protocol

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CINC-1, terbutaline and mouse monoclonal ANTI-FLAG BioM2 antibody (Ab) were purchased from Sigma (St. Louis, MO, USA); ADRB2 polyclonal Ab, FITC conjugated (H-20) was purchased from Bioss (Woburn, MA, USA); GAPDH monoclonal Ab (14C10) was purchased from Cell Signaling; PIK-90 was purchased from Cayman Chemical (Ann Arbor, MI, USA); protein A–Sepharose beads were purchased from Pierce (Rockford, IL, USA); PDE 4D polyclonal Ab (14613) was purchased from Abcam (Cambridge, MA, USA); rolipram was purchased from MP Biomedicals (Waltham, MA, USA); Odyssey Blocking Buffer, IRDye® 680RD and 800CW Donkey anti-Rabbit IgG Abs and IRDye® 680 RD and 800CW Donkey anti-Mouse IgG Abs were purchased from LI-COR (Lincoln, NE, USA); Primocin was purchased from InvivoGen (San Diego, CA, USA); Precision Red Advanced Protein Assay was purchased from Cytoskeleton (Denver, CO, USA); all other reagents were purchased from Fisher Scientific (Waltham, MA, USA).
Rich T.C., Leavesley S.J., Brandon A.P., Evans C.A., Raju S.V, & Wagener B.M. (2021). Phosphodiesterase 4 Mediates Interleukin-8-Induced Heterologous Desensitization of the β2-Adrenergic Receptor. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 35(10), e21946.
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4

Antibody Immunoblotting Protocols

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The following primary antibodies were used for immunoblotting: mouse polyclonal anti-DDAH1 (Thermo Fisher Scientific, H00023576-BO1P), rabbit polyclonal Anti-CYR61/CCN1 (Abcam, ab10760), mouse monoclonal anti-MVP (Thermo Fisher Scientific, MA5-13871), mouse monoclonal anti-ISG15 (F-9, sc-166755, Santa Cruz Biotechnology), rabbit polyclonal anti-RNF213 (HPA003347, Merck), rabbit polyclonal anti-tubulin-α antibody (#ab18251, Abcam), mouse monoclonal anti-FLAG-tag (M2, #F3165, Merck). Aforementioned primary antibodies were revealed using goat polyclonal anti-mouse IgG (IRDye® 800CW, Li-COR), goat polyclonal anti-rabbit-IgG (IRDye® 800CW, Li-COR), goat polyclonal anti-mouse-IgG (IRDye® 680RD, Li-COR) or goat polyclonal anti-rabbit-IgG (IRDye®680RD, Li-COR), except for anti-RSV serum which was revealed with secondary anti-goat (#sc-2020, Santa Cruz biotechnology).
Martina L., Asselman C., Thery F., Boucher K., Delhaye L., Maia T.M., Dermaut B., Eyckerman S, & Impens F. (2021). Proteome Profiling of RNF213 Depleted Cells Reveals Nitric Oxide Regulator DDAH1 Antilisterial Activity. Frontiers in Cellular and Infection Microbiology, 11, 735416.
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5

Protein Extraction and Western Blot Analysis

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Total protein was extracted with radioimmunoprecipitation assay buffer (150 mM NaCl, 1% TritionX-100, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulfate [SDS], 50 mM Tris pH 8.0) supplemented with Complete EDTA-free protease inhibitors (Roche), 5 mM NaF, and 1 mM Na3VO4. Nuclear extraction was performed by suspending cells in a hypotonic buffer (10 mM HEPES-KOH pH 7.9, 1.5 mM MgCl2, 10 mM KCl, 1 mM DTT, 1x Halt Protease Inhibitor [ThermoFisher; Cat # 78430]). After dounce homogenization and centrifugation, the resulting nuclear pellets were suspended in a hypertonic buffer (20 mM HEPES-KOH pH 7.9, 25% Glycerol, 1.5 mM MgCl2, 0.6 M KCl, 0.2 mM EDTA, 1 mM DTT, 1x Halt Protease Inhibitor). Protein was separated on an SDS polyacrylamide gel electrophoresis and transferred to polyvinylidene fluoride membrane. Primary antibody incubation was performed overnight at 4C. Secondary goat anti-rabbit (IRDye 680RD or IRDye 800CW, LI-COR, Cat # 92568071 [RRID: AB_2721181] or Cat # 926–32211 [RRID: AB_621842]; 1:20,000) and goat anti-mouse (IRDye 680RD or IRDye 800CW, LI-COR, Cat # 926–68070 [RRID: AB_10956588] or Cat# 925–32210 [RRID: AB_2687825], 1:20,000) antibodies were applied for 1 hour at room temperature. Blots were visualized using the Licor Odyssey Imaging System and ImageStudio software (V4).
McMellen A., Yamamoto T.M., Qamar L., Sanders B.E., Nguyen L.L., Chavez D.O., Bapat J., Berning A., Post M.D., Johnson J., Behbakht K., Nurmemmedov E., Chuong E.B, & Bitler B.G. (2023). ATF6-mediated signaling contributes to PARP Inhibitor Resistance in Ovarian Cancer. Molecular cancer research : MCR, 21(1), 3-13.
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6

Western Blot Analysis of Protein Expression

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Protein was extracted with RIPA buffer (150 mM NaCl, 1%
TritonX‐100, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulfate [SDS],
50 mM Tris pH 8.0) supplemented with Complete EDTA‐free protease
inhibitors (Roche), Sodium Fluoride (10 mmol/L), and Sodium Orthovanadate (1
mmol/L). Protein was separated on an SDS-PAGE and transferred to polyvinylidene
fluoride membrane. Primary antibody incubation was performed overnight at
4°C. Secondary goat anti‐rabbit (IRDye 680RD or IRDye 800CW,
LI‐COR, Cat # 92568071 or Cat # 926‐32211; 1:20,000) and goat
anti‐mouse (IRDye 680RD or IRDye 800CW, LI‐COR, Cat #
926‐68070 or Cat#925‐32210, 1:20,000) antibodies were applied for
1 hour at room temperature. Blots were visualized using the LI-COR Odyssey
Imaging System. For development of Western blots using horseradish peroxidase
(HRP) detection, antibody was diluted in 5% BSA/TBST. The secondary antibody
(Cell signaling, Cat# 7076S or #7074P2) was used at concentration of 1:3000 and
HRP enzymatic activity was visualized with SuperSignal West Femto Maximum
Sensitivity Substrate (Thermo Scientific) in G:box (SYNGENE). Densitometry of
immunoblots was completed in ImageJ (NIH) by normalizing the integrated density
of the experimental antibody with the loading control.
Sanders B.E., Yamamoto T.M., McMellen A., Woodruff E.R., Berning A., Post M.D, & Bitler B.G. (2022). Targeting DUSP activity as a treatment for high grade serous ovarian carcinoma. Molecular cancer therapeutics, 21(8), 1285-1295.
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7

Comprehensive Protein Extraction and Western Blot

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Total protein was extracted with radioimmunoprecipitation assay buffer (150 mM NaCl, 1% TritionX-100, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulfate [SDS], 50 mM Tris pH 8.0) supplemented with Complete EDTA-free protease inhibitors (Roche, Cat. #4693132001), 5mM NaF, and 1mM Na3VO4. Nuclear extraction was performed by suspending cells in a hypotonic buffer (10mM HEPES-KOH pH 7.9, 1.5mM MgCl2, 10mM KCl, 1mM DTT, 1x Halt Protease Inhibitor). After dounce homogenization and centrifugation, the resulting nuclear pellets were suspended in a hypertonic buffer (20mM HEPES-KOH pH 7.9, 25% Glycerol, 1.5mM MgCl2, 0.6M KCl, 0.2mM EDTA, 1mM DTT, 1x Halt Protease Inhibitor). Protein was separated on an SDS polyacrylamide gel electrophoresis and transferred to polyvinylidene fluoride membrane. Primary antibody incubation was performed overnight at 4C. Secondary goat anti-rabbit (IRDye 680RD or IRDye 800CW, LI-COR, Cat. #92568071; RRID: AB_2721181 or Cat. #926-32211; RRID: AB_621842; 1:20,000) and goat anti-mouse (IRDye 680RD or IRDye 800CW, LI-COR, Cat. # 926-68070; RRID: AB_10956588 or Cat# 925-32210; RRID: AB_2687825; 1:20,000) antibodies were applied for 1 hour at room temperature. Blots were visualized using the Licor Odyssey Imaging System and ImageStudio software (V4).
Nguyen L.L., Watson Z.L., Ortega R., Woodruff E.R., Jordan K.R., Iwanaga R., Yamamoto T.M., Bailey C.A., Jeong A.D., Guntupalli S.R., Behbakht K., Gbaja V., Arnoult N., Chuong E.B, & Bitler B.G. (2023). Combinatory EHMT and PARP inhibition induces an interferon response and a CD8 T cell-dependent tumor regression in PARP inhibitor-resistant models. bioRxiv.
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8

Modulation of T-cell signaling by SLAMF6

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Anti-CD3 (UCHT1) and PE conjugated anti-SLAMF6 (292811) were purchased from R&D Systems. Goat-anti-mouse IgG (Poly4053), Alexa Fluor 647 conjugated anti-ERK (4B11B69), phospho Thr202/Tyr204, Alexa Fluor 647 conjugated IgG2b iso-type control (MOPC-173), and anti-SLAMF6 (NT-7) were purchased from BioLegend. SLAMF6 (HA12OC3104) was purchased from Sino Biologics Inc. PE conjugated IgG2a iso-type control (eBR2a) was purchased from eBioscience. Western blot antibodies for pERK (E10) Thr202/Tyr204, pZAP70 (Y352) Tyr309, pAKT (L32A4) Thr308 and pSRC (Y527) were purchased from Cell Signaling. Anti-beta actin (AC-15) was purchased from Invitrogen. Western blot secondary antibodies IRDye 680RD Goat anti-mouse IgG, IRDye 800CW Goat anti-mouse IgG and IRDye 680RD Goat anti-rabbit IgG were purchased from Licor. Anti-SLAMF6 (NT-7) Fab fragments were generated using the Pierce Fab Preparation Kit obtained from Thermo Fisher. Cells were stimulated with the following soluble antibodies: anti-CD3/anti-mouse IgG (3.25 μg/mL), anti-SLAMF6 (3.25 μg/mL), anti-mouse IgG (1.63 μg/mL), anti-SLAMF6 Fab (3.25 μg/mL) anti-mouse IgG (1.63 μg/mL). Cells were stimulated with the following immobilized antibodies: anti-CD3/anti-mouse IgG (1.5 μg/mL), anti-SLAMF6 (5 μg/mL).
Dragovich M.A., Adam K., Strazza M., Tocheva A.S., Peled M, & Mor A. (2019). SLAMF6 clustering is required to augment T cell activation. PLoS ONE, 14(6), e0218109.
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9

Western Blot Protein Analysis

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Protein was extracted with RIPA buffer (150 mM NaCl, 1% TritonX-100, 0.5% sodium deoxycholate, 0.1% SDS, 50 mM Tris pH 8.0) supplemented with Complete EDTA-free protease inhibitors (Roche), NaF and Na3VO4. Protein was separated on a SDS-PAGE and transferred to PVDF membrane. Primary antibody incubation was performed overnight at 4°C. Secondary goat anti-rabbit (IRDye 680RD or IRDye 800CW, LI-COR, Cat # 92568071 or Cat # 926-32211, 1:20,000) and goat anti-mouse (IRDye 680RD or IRDye 800CW, LI-COR, Cat # 926-68070 or Cat# 925-32210, 1:20,000) antibodies were applied for one hour at room temperature. Blots were visualized using the Licor Odyssey Imaging System. For WNT3A immunoblotting, antibodies were diluted in 5% milk/TBST (50 mM Tris pH7.5, 150 mM NaCl, 0.1% Tween20) blocking buffer and HRP chemiluminescent signal was detected with SuperSignal West Femto (Thermo scientific) and visualized using the G:Box (SYNGENE).
Yamamoto T.M., McMellen A., Watson Z.L., Aguilera J., Ferguson R., Nurmemmedov E., Thakar T., Moldovan G.L., Kim H., Cittelly D.M., Joglar A.M., Brennecke E.P., Wilson H., Behbakht K., Sikora M.J, & Bitler B.G. (2019). Activation of Wnt signaling promotes olaparib resistant ovarian cancer. Molecular carcinogenesis, 58(10), 1770-1782.
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10

Western Blot Analysis of Proteins

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Proteins obtained in total cell lysates or by immunoprecipitation were denatured in Laemmli buffer at 95°C for 5 min, separated by SDS-PAGE, and then transferred to nitrocellulose membrane and revealed with specific primary antibodies, followed by the addition of IRdye secondary antibodies (goat anti-Rat IRDye 680RD, donkey anti-mouse IRDye 680RD, and donkey anti-rabbit IRDye 800CW; Li-Cor Biosciences), and analyzed by imaging with an Odyssey infrared imaging system CLx (Li-Cor Biosciences).
Boson B., Mialon C., Schichl K., Denolly S, & Cosset F.L. (2022). Nup98 Is Subverted from Annulate Lamellae by Hepatitis C Virus Core Protein to Foster Viral Assembly. mBio, 13(2), e02923-21.
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