Irdye 680rd donkey anti rabbit igg

Manufactured by LI COR

Sourced in United States

IRDye 680RD Donkey anti-Rabbit IgG is a secondary antibody conjugated with the near-infrared fluorescent dye IRDye 680RD. It is designed to detect and quantify rabbit primary antibodies in various immunoassay applications.

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Lab products found in correlation

Irdye 800cw donkey anti mouse igg, by LI COR (20 mentions) Odyssey clx imaging system, by LI COR (5 mentions) Odyssey clx, by LI COR (4 mentions) Odyssey blocking buffer, by LI COR (4 mentions) Irdye 800cw goat anti mouse igg, by LI COR (3 mentions) Irdye 800cw donkey anti rabbit igg, by LI COR (3 mentions) Protease inhibitor cocktail, by Roche (3 mentions) Odyssey imaging system, by LI COR (2 mentions) Intercept blocking buffer, by LI COR (2 mentions) Immobilon fl pvdf membrane, by Merck Group (2 mentions) Image studio software, by LI COR (2 mentions) Irdye 680rd goat anti mouse igg, by LI COR (2 mentions) Nbp1 71915, by Novus Biologicals (2 mentions) Pi3k p85α, by Cell Signaling Technology (2 mentions) Erk1 2, by Cell Signaling Technology (2 mentions) Nf κb p65, by Cell Signaling Technology (2 mentions) Stat3, by Cell Signaling Technology (2 mentions) β tubulin, by Santa Cruz Biotechnology (2 mentions) Precast protein gel, by Bio-Rad (2 mentions) β actin, by Thermo Fisher Scientific (2 mentions)

Spelling variants of this product

IRDye 680RD (357 citations) IRDye 680RD donkey anti-rabbit (39 citations) Anti-rabbit IRDye 680RD (17 citations) IRDye 680RD anti-rabbit IgG (8 citations) IRDye 680RD Donkey anti-Rabbit IgG secondary antibody (7 citations) IRDye 680RD donkey anti-rabbit IgG (H (3 citations) Donkey anti‐Rabbit IRDye (2 citations)

36 protocols using irdye 680rd donkey anti rabbit igg

FLI1 Expression Analysis in Dox-Treated Cells

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Cell lysates from both Dox-treated and untreated cells were subjected to western blotting using a 1:1000 dilution of rabbit anti-human FLI1 (Thermo Scientific, #PA1-21023), or 1:2500 dilution of mouse anti-β-tubulin (Sigma-Aldrich, T4026) as primary antibodies, followed by treatment with a 1:100000 dilution of IRDye 680RD donkey anti-mouse IgG secondary antibody (LI-COR, #926–68072) and IRDye 680RD donkey anti-Rabbit IgG (LI-COR, #926–68073).
The fractionated cell samples obtained from Dox-treated and untreated stable cells were subjected to western blotting using a 1:1000 dilution of rabbit anti-human FLI1 (Thermo Scientific, #PA1-21023), followed by treatment with a 1:10,000 dilution of IRDye 680RD donkey anti-Rabbit IgG (LI-COR, #926–68073). The western blot of the loading controls was treated with a 1:1000 dilution of rabbit anti-Histone H2A (Abcam, #ab18255) and a 1:2000 dilution of mouse anti-β-tubulin (Sigma-Aldrich, T4026) antibodies. All images of western blots were captured using the LI-COR Odyssey Imaging System.

Park H., Kim H., Hassebroek V., Azuma Y., Slawson C, & Azuma M. (2020). Chromosomal localization of Ewing sarcoma EWSR1/FLI1 protein promotes the induction of aneuploidy. The Journal of Biological Chemistry, 296, 100164.

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Immunoblotting and Subcellular Localization of Mycobacterial Proteins

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A total of 10 µg of WCL was used for immunoblotting or subcellular localization for detection of TNT, EsxF, and EsxE. RNAP was used as a loading control for WCLs and as a lysis control for in vitro secretion assays, GlpX was used as a water-soluble compartment control, and MctB was used as a membrane compartment control for subcellular localization. A total of 50 µg was used to detect EsxF or EsxE due to the weakness of the polyclonal antibodies. LpqH was used as a secreted protein control as it is found in extracellular vesicles of Mtb. Samples were prepared with Laemmli protein loading dye and run on a 10% SDS–PAGE gel and transferred overnight with a PVDF membrane. Blots were blocked with Odyssey blocking buffer in TBST (LI-COR Biosciences) with two washes with TBST between detection steps. Primary antibody detection was performed using rabbit polyclonal anti-TNT (1:1000), rabbit polyclonal anti-EsxF (1:50), rabbit polyclonal anti-EsxE (1:50), mouse monoclonal anti-RNAP (1:1000, BioLegend), mouse monoclonal anti-LpqH (1:1000, BEI Resources), rabbit polyclonal anti-GlpX (1:1000, BEI Resources), and anti-MctB (1:1000). Secondary detection was performed with donkey anti-rabbit IgG-IRDye 680RD or goat anti-mouse IgG-IRDye 800CW, which were obtained from LI-COR biosciences.

Tak U., Dokland T, & Niederweis M. (2021). Pore-forming Esx proteins mediate toxin secretion by Mycobacterium tuberculosis. Nature Communications, 12, 394.

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EV Protein Characterization by SDS-PAGE and Immunoblotting

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For SDS‐PAGE, a volume of EVs corresponding to 1.5 μg of total protein was prepared in Laemmli sample buffer (BioRad 1610747) containing β‐mercaptoethanol and heated at 95°C for 5 minutes. Proteins were separated through a 4%–12% NuPAGE Bis‐Tris gel (Invitrogen), transferred to an Immobilon‐FL PVDF membrane (Millipore), and detected as described previously (Yang et al., 2011 (link)). After blocking with Intercept blocking buffer (Licor Biosciences), the following antibodies were used: mouse anti‐calnexin (BD Biosciences 610523, 1:500), rabbit anti‐GM130 (Abcam ab52649, 1:500), mouse anti‐cytochrome c (BD Biosciences 556433, 1:1000), rabbit anti‐CD9 (Cell Signaling Technologies 13174, 1:1000), mouse anti‐TSG101 (BD Biosciences 612696, 1:500), and mouse anti‐THP (Santa Cruz Biotechnology sc‐271022, 1:1000), rabbit anti‐beta‐actin (Invitrogen, PA1‐16889, 1:5000), donkey anti‐mouse IgG IRDye 800CW (LiCor 925–32212, 1:10,000), and donkey anti‐rabbit IgG IRDye 680RD (LiCor 926–68073, 1:10,000).

Correll V.L., Otto J.J., Risi C.M., Main B.P., Boutros P.C., Kislinger T., Galkin V.E., Nyalwidhe J.O., Semmes O.J, & Yang L. (2022). Optimization of small extracellular vesicle isolation from expressed prostatic secretions in urine for in‐depth proteomic analysis. Journal of Extracellular Vesicles, 11(2), e12184.

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Fluorescent Western Blotting of PDGFRα

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HEK293T cells were seeded (7.5 × 10⁶ cells) into 90 mm dishes and transfected with wild-type or mutated PDGFRα expression vectors using Lipofectamine 3000. Forty-eight h after, the transfected cells were lysed using the Mem-PER Plus membrane protein extraction kit (Thermo Fisher Scientific), and proteins were visualized via fluorescent western blotting. Briefly, proteins were resolved by SDS-PAGE and transferred onto PVDF membranes. Next, membranes were blocked and incubated with antibodies using the protocol described above. The anti-PDGFRα (#5214, Cell Signaling Technology) and anti-phosphotyrosine (4G10) antibodies were used as the primary antibodies, and IRDye 680RD donkey anti-rabbit IgG and IRDye 800CW donkey anti-mouse IgG (LI-COR, Lincoln, NE, USA) were used as the secondary antibodies. Fluorescence detection and quantification were performed using Odyssey CLx (LI-COR).

Hamada T., Akahane T., Yokoyama S., Higa N., Kirishima M., Matsuo K., Shimokawa M., Yoshimoto K, & Tanimoto A. (2022). An oncogenic splice variant of PDGFRα in adult glioblastoma as a therapeutic target for selective CDK4/6 inhibitors. Scientific Reports, 12, 1275.

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Quantitative Western Blot Analysis of BDNF and TrkB

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HIP and PFC samples (20 µg total protein) were separated on 10% criterion TGX gels (Bio-Rad) and transferred to nitrocellulose membranes. The samples were then blocked in odyssey blocking buffer (LI-COR, Lincoln, NE, USA) and probed with the primary antibodies: rabbit anti-BDNF (Sigma AV41970; 1:2000), mouse anti-β-actin (926-42212, LI-COR; 1:3000), goat anti-TrkB (AF1494, R&D Systems, Minneapolis, MN, USA; 1:500), and rabbit anti-TrkB(Y817) (ab81288, Abcam, Cambridge, UK; 1:1000) overnight at 4 °C. This was followed by incubation with the appropriate IRDye conjugated secondary antibody for 1 h at RT: IRDye 800CW donkey anti-goat IgG, IRDye 680RD donkey anti-rabbit IgG, IRDye 800CW goat anti-rabbit IgG, or IRDye 680RD goat anti-mouse IgG, all in 1:15,000 dilution (LI-COR). Infrared signals were detected using the Odyssey CLx infrared imaging system (LI-COR, and bands were quantified using Image Studio software (LI-COR).

Virtuoso A., Tveden-Nyborg P., Schou-Pedersen A.M., Lykkesfeldt J., Müller H.K., Elfving B, & Sørensen D.B. (2021). A Long-Term Energy-Rich Diet Increases Prefrontal BDNF in Sprague-Dawley Rats. Nutrients, 14(1), 126.

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STING Protein Expression Regulation

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RAW 264.7 cells (1 × 10⁶ cells) were cultured, then underwent DMXAA treatment with and without RRBE for 3 h. Cells were lysed in buffer C (final concentration at 150 mM of NaCl, 5 mM of EDTA pH 8, 1% of Triton-X100 and 10 mM of Tris-HCL pH 7.4). Protein lysates were homogenized and centrifuged at 12,000× g, 15 min at 4 °C. The proteins were collected in the supernatants. Then, total protein was examined with a BCA assay (Thermo Fisher Scientific, Waltham, MA, USA). Next, 15 µg of total protein was boiled in Laemmli buffer at 95 °C for 5 min. The equal proteins were separated on a 12.5% SDS-polyacrylamide gel. Then, nitrocellulose membranes were used for transferring the proteins and blocking the unspecific binding of the antibodies with 5% BSA for 1 h. The membranes were probed with STING antibody (clone: D2P2F cat: 13647, 1:2000) (Cell Signaling, Danvers, MA, USA) and incubated at 4 °C overnight. The membranes were washed with wash buffer and probed with the fluorescent secondary antibody IRDye^® 680RD donkey anti-rabbit IgG (1:10,000) (LI-COR, Lincoln, NE, USA) at room temperature for 1 h. Membranes were determined by protein signals using ODYSSEY CLx (LI-COR, Lincoln, NE, USA).

Onsa-Ard A., Thongboontho R., Munkong N., Phromnoi K., Ontawong A., Pengnet S, & Thim-Uam A. (2022). Anti-Inflammatory Effects of Red Rice Bran Extract Ameliorate Type I Interferon Production via STING Pathway. Foods, 11(11), 1622.

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Protein Detection via SDS-PAGE and Western Blotting

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Proteins were detected via SDS-PAGE and Western blotting as described in Horos et al.^{4 (link)}. Antibodies used were directed against Csde1 (NBP1-71915, Novus Biological), Actin (A3853, Sigma-Aldrich) and alpha Tubulin (ab4074, Abcam). Fluorescently labeled secondary antibodies for visualization with Odyssey were IRDye 680RD Donkey anti-Rabbit IgG (926–68073, Licor) and IRDye 800CW Donkey anti-Mouse IgG (925–32212, Licor).

Moore K.S., Yagci N., van Alphen F., Paolini N.A., Horos R., Held N.M., Houtkooper R.H., van den Akker E., Meijer A.B., ‘t Hoen P.A, & von Lindern M. (2018). Csde1 binds transcripts involved in protein homeostasis and controls their expression in an erythroid cell line. Scientific Reports, 8, 2628.

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Immunoblotting Analysis of Cellular Signaling

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Whole cell lysates were prepared and the immunoblotting was done as previously described [3 (link)] in Triton X 100-containing lysis buffer (50 mM Tris–HCl, pH 7.5, 10% glycerol, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 50 mM NaF). For Western blotting, the following antibodies were used: pAKT (S473, #6942), AKT (#9272), Bcl-xL (#2764), p ERK1/2 (#9101), MEK1/2 (#4694), pMEK1/2 (#9121), ERK1/2 (#4696), NF-κB p65 (#6956), pNF-κB p65 (#3033), PARP (#9532), PI3K p85α (#13,666), pPI3K p85/p55 (#4228), PTK7 (#25618), Rac-1 (#4651), RhoA (#2117), ROR1 (#16,540), ROR2 (#88,639), Src (#2109), STAT3 (#9139), pSTAT3 (#9145) from Cell Signaling Technology (CST, Danvers, MA, USA); anti-pTYR 4G10 (#05–321) from Merck Millipore (Burlington, MA, USA); β-tubulin (#sc-166729) from Santa Cruz Biotechnology (Dallas, TX, USA); HA (#901,513) from BioLegend (San Diego, CA, USA). As secondary antibodies, IRDye^® 800CW Donkey anti-Mouse IgG or IRDye^® 680RD Donkey anti-Rabbit IgG (LI-COR, Lincoln, NE, USA) were used at 1:10 000 dilution.

Raivola J., Dini A., Salokas K., Karvonen H., Niininen W., Piki E., Varjosalo M, & Ungureanu D. (2022). New insights into the molecular mechanisms of ROR1, ROR2, and PTK7 signaling from the proteomics and pharmacological modulation of ROR1 interactome. Cellular and Molecular Life Sciences, 79(5), 276.

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Quantitative Western Blot Analysis Protocol

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The protein extracts from the cells (40 μg) were subjected to
electrophoresis in SDS-PAGE on 4–15% precast protein gels (Bio-Rad,
Hercules, CA), separated and transferred to polyvinylidene fluoride (PVDF, 0.2
μm pore size) membranes. The membranes were probed with primary
antibodies to the androgen receptor (GeneTex, Irvine, CA) and β-actin
(Thermo Fisher Scientific) overnight at 4 °C. After washing, the
membranes were incubated with the secondary antibodies IRDye® 680RD
donkey anti-rabbit IgG and IRDye® 800CW donkey anti-mouse IgG (Li-COR,
Lincoln, NE). The bands were visualized by scanning the fluorescence signal via
the Odyssey® CLx imaging system (LI-COR). Then the photographic images
with blots were scanned and quantified by the software of the system for the
comparison of the fluorescence value of the blots.

Guan J., Guo H., Tang T., Wang Y., Wei Y., Seth P., Li Y., Dehm S.M., Ruoslahti E, & Pang H.B. (2021). iRGD-liposomes enhance tumor delivery and therapeutic efficacy of antisense oligonucleotide drugs against primary prostate cancer and bone metastasis. Advanced functional materials, 31(24), 2100478.

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Protein Expression Analysis by Western Blot

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Cells were lysed with RIPA lysis buffer (Thermo Scientific #89900) supplemented with a 1× protease and phosphatase inhibitor mixture (Thermo Scientific #78440), rotated at 4 °C for 30 min, then centrifuged at 13,000 × g for 30 min. Protein concentration was quantified using the bicinchoninic acid assay (Thermo Scientific #23225). Around 40 μg of protein was run on a 4–12% Bis-Tris gradient gel (Invitrogen #NP0335), then transferred onto a nitrocellulose membrane. The following primary antibodies were used for immunoblotting: rabbit anti-p107 (Santa Cruz #sc-318, 1:500), rabbit anti-p130 (Santa Cruz #sc-317, 1:500), and rabbit anti-β-actin (Cell Signaling Technology #4970, 1:10,000). Primary antibodies were detected with the following fluorescent secondary antibodies: IRDye 680RD donkey anti-rabbit IgG (LI-COR #926–68073, 1:10,000), IRDye 800CW donkey anti-rabbit IgG (LI-COR # 926–32,213, 1:10,000). Immunoblots were imaged using the LI-COR Odyssey infrared imager and quantified using Image Studio (LI-COR).

Ng S.R., Rideout WM I.I.I., Akama-Garren E.H., Bhutkar A., Mercer K.L., Schenkel J.M., Bronson R.T, & Jacks T. (2019). CRISPR-mediated modeling and functional validation of candidate tumor suppressor genes in small cell lung cancer. Proceedings of the National Academy of Sciences of the United States of America, 117(1), 513-521.

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