Goat anti rat af488

Manufactured by Thermo Fisher Scientific

Goat anti-rat AF488 is a fluorescent-labeled secondary antibody used in immunofluorescence applications. It is designed to detect and bind to primary antibodies raised in rat. The AF488 fluorophore attached to the antibody emits green fluorescence when excited, allowing for visualization and detection of target proteins or cells.

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Lab products found in correlation

Lsm 880 confocal microscope, by Zeiss (3 mentions) Ho 1 antibody, by Enzo Life Sciences (2 mentions) Normal goat serum (ngs), by Thomas Scientific (2 mentions) Molecular probes prolong diamond antifade mountant, by Thermo Fisher Scientific (2 mentions) Ix83p2f microscope, by Olympus (2 mentions) Axio imager a1 microscope, by Zeiss (2 mentions) Axiocam mrc5, by Zeiss (2 mentions) Goat anti mouse af647, by Thermo Fisher Scientific (2 mentions) Decloaking chamber, by Biocare Medical (2 mentions) Goat anti chicken af488, by Thermo Fisher Scientific (2 mentions) Goat anti rabbit af594, by Thermo Fisher Scientific (2 mentions) Rnascope multiplex fluorescent reagent kit v2, by Advanced Cell Diagnostics (2 mentions) Fluoview fv1000, by Olympus (1 mentions) Dextran af488, by Thermo Fisher Scientific (1 mentions) Wga af647, by Thermo Fisher Scientific (1 mentions) Donkey anti goat af555, by Thermo Fisher Scientific (1 mentions) Chicken anti rabbit af488, by Thermo Fisher Scientific (1 mentions) Dq green bsa, by Thermo Fisher Scientific (1 mentions) Lamp1, by BD (1 mentions) Clone 6f11, by Leica (1 mentions)

Close spelling variants of this product

Anti-rat AF488 (4 citations) Goat anti-rat IgG-AF488 (2 citations)

13 protocols using goat anti rat af488

Immunohistochemistry for HO-1 and F4/80 in Paraffin Sections

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Paraffin embedded tissues were deparaffinized using two 100% xylene washes for 5 minutes followed by sequential 3 minute washes in 1:1 xylene:Ethanol, 100%, 95%, 70%, and 50% ethanol. Tissues were then rinsed and placed in a staining box with Tris-EDTA buffer pH 9.0 (10mM Tris Base, 1mM EDTA solution, 0.05% TWEEN®20). Heat induced epitope retrieval was performed at 95°C in a pressure cooker for 25 minutes. After rinsing in 1x TBST buffer (Tris-buffered saline, 0.2% TWEEN®20) slides were blocked with 10% normal goat serum (Thomas Scientific) in PBS for 1 hour. After 1x TBST rinse, staining was performed with rabbit polyclonal HO-1 antibody (Enzo Life Sciences), and rat monoclonal [CI:A3–1] F4/80 antibody (Abcam) 4°C overnight in a humidified chamber. After rinsing, secondary antibodies goat anti-rabbit AF-555 and goat anti-rat AF-488 (Invitrogen), were applied for 2 hours at room temperature in a humidified chamber. Slides were rinsed then sealed with Molecular Probes™ ProLong™ Diamond Antifade Mountant with or without DAPI (Fischer Scientific). Imaging was done on an Olympus IX83P2F microscope equipped with a DP80 camera. Additional images were obtained on an AxioImager A1 microscope (Zeiss) equipped with an AxioCam MRc5.

Schaefer R.E., Callahan R.C., Atif S.M., Orlicky D.J., Cartwright I.M., Fontenot A.P., Colgan S.P, & Onyiah J.C. (2021). Disruption of monocyte-macrophage differentiation and trafficking by a heme analog during active inflammation. Mucosal immunology, 15(2), 244-256.

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Immunocytochemical Staining of Transfected HEK 293T Cells

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Transfected HEK 293T cells were seeded on poly-L-lysine-coated coverslips for staining. Cells were fixed with 2% paraformaldehyde (PFA), permeabilized in PBS containing 0.1% Triton X-100, and blocked with 20% goat serum in PBS. Cells were incubated with rat monoclonal anti-PTX3 (MNB4, Abcam) or mouse monoclonal anti-alphavirus (3581, Santa Cruz) primary antibody in PBS, followed by goat anti-rat AF488 (Invitrogen) or goat anti-mouse AF647 (Invitrogen) secondary antibody. Cells were washed, mounted, and examined with a confocal laser-scanning microscope (Fluoview FV 1000, Olympus) at 60x magnification. Images were collected and processed using FV1000-ASW software.

Foo S.S., Chen W., Taylor A., Sheng K.C., Yu X., Teng T.S., Reading P.C., Blanchard H., Garlanda C., Mantovani A., Ng L.F., Herrero L.J, & Mahalingam S. (2015). Role of Pentraxin 3 in Shaping Arthritogenic Alphaviral Disease: From Enhanced Viral Replication to Immunomodulation. PLoS Pathogens, 11(2), e1004649.

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Embryonic Vascular and Hematopoietic Staining

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Embryos were stained using a rat anti-mouse CD31 antibody (1:500) (BD biosciences; 553,370) and a goat anti-rat AF555 secondary antibody (1:1000) (Invitrogen) as previously described (Yokomizo et al., 2012 (link)). Z-stack images were taken using a two-photon confocal microscope with a 5 × objective (Leica). E10.5 embryo sections were stained as previously described (Thambyrajah et al., 2016 (link)) using a goat anti-SOX7 antibody (1:200) (R&D systems; AF2766) and a donkey anti-goat AF647 antibody (1:2000) (Invitrogen). Subsequently, embryos were stained with a rat anti-cKit antibody (1:1000) (BD biosciences; 553,868), and a rabbit anti-pan-RUNX antibody (1:1000) (Abcam; ab92336) before staining with a goat anti-rat AF488 and a goat anti-rabbit antibody (both 1:2000) (both Invitrogen). Yolk sacs were isolated and flat mounted with DAPI as previously described (Frame et al., 2016 (link)) before imaging. Specific labelling of primary antibodies was determined by comparison with no primary antibody stained controls.

Lilly A.J., Mazan A., Scott D.A., Lacaud G, & Kouskoff V. (2017). SOX7 expression is critically required in FLK1-expressing cells for vasculogenesis and angiogenesis during mouse embryonic development. Mechanisms of Development, 146, 31-41.

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Fluorescent Markers for Organelle Tracking

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Primary antibodies included the following: EEA1 (Santa Cruz Biotechnology, sc-6415), Rab7 (Abcam, ab 137029), Lamp1 (BD Pharmingen, 1D4B), GFP (Life Technologies, A11122). Secondary antibodies included the following: donkey anti-goat–AF555 (Invitrogen, A21432), goat anti-rat–AF488 (Invitrogen A11006), chicken anti-rabbit–AF488 (Invitrogen, A21441).
Other reagents included the following: AF594/Fab (Jackson ImmunoResearch Laboratories Inc., 309-586-003), A488 phalloidin (Life Technologies, A12379), AF555 phalloidin (Life Technologies, A34055), DQ Green BSA (Life Technologies, D12050), Tf-AF647 (Life Technologies, T23366), dextran-AF488 (Life Technologies, ID7178), WGA-AF647 (Life Technologies, W32466).

Piperno G.M., Naseem A., Silvestrelli G., Amadio R., Caronni N., Cervantes-Luevano K.E., Liv N., Klumperman J., Colliva A., Ali H., Graziano F., Benaroch P., Haecker H., Hanna R.N, & Benvenuti F. (2020). Wiskott-Aldrich syndrome protein restricts cGAS/STING activation by dsDNA immune complexes. JCI Insight, 5(17), e132857.

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Immunohistochemistry for HO-1 and F4/80 in Paraffin Sections

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Multimarker Immunohistochemistry of Mammary Gland

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Expression of BrdU (1:100; clone BU1/75, Abcam), ER (1:100; clone 6F11, Leica Biosystems), Aldh1a3 (1:200; clone HPA046271, Sigma), Krt 5 (1:500; polyclonal, Abcam), and/or Krt 14 (1:1000; polyclonal, Covance) in intact mammary glands was done by fixing freshly isolated glands in 4% PFA overnight and processing the tissue into paraffin. Tissue blocks were sectioned at 4 µm, deparaffinised in xylene, gradually rehydrated in descending concentrations of ethanol, and subsequently treated in Borg Decloaker antigen retrieval solution (pH 6) for 30 min at 121 °C and 10 s at 90 °C using a Decloaking chamber (Biocare Medical). The samples were preblocked in PBS with 1% BSA and 0.1% Tween 20, and then incubated with primary antibodies overnight at 4 °C. The secondary antibodies were goat anti-mouse AF647, goat anti-rabbit Cy3, goat anti-rat AF488 and/or goat anti-chicken AF488 (Invitrogen). No primary antibody was used as a control. Sections were mounted with ProLong Gold antifade with DAPI. Tissue sections were imaged using the Olympus BX50 microscope (Olympus).

Shehata M., Kim H., Vellanki R., Waterhouse P.D., Mahendralingam M., Casey A.E., Koritzinsky M, & Khokha R. (2019). Identifying the murine mammary cell target of metformin exposure. Communications Biology, 2, 192.

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Immunohistochemical Analysis of Mammary Gland

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Freshly isolated intact mammary glands were fixed in 4% PFA and processed into paraffin. A total of 4 µm sections were deparaffinised in xylene, gradually rehydrated in descending concentrations of ethanol, and subsequently treated in Borg Decloaker antigen retrieval solution (pH 6) for 30 min at 121 °C and 10 s at 90 °C using a Decloaking chamber (Biocare Medical). The samples were preblocked in PBS with 1% BSA and 0.1% Tween 20, before overnight incubation at 4 °C with primary antibodies: anti-BrdU (clone BU1/75, Abcam), anti-BrdU (Clone B44, BD Biosciences), anti-ERα (6F11, Novocastra), anti-PR (H190, Santa Cruz), anti-Keratin 5 (polyclonal, Abcam), anti-DsRed (polyclonal, Clontech), anti-GFP (polyclonal, Abcam), anti-Ki-67 (clone: SolA15, Thermo Fisher). The secondary antibodies were goat anti-mouse AF647, goat anti-rabbit Cy3 (Jackson Labs), goat anti-rat AF488 and/or goat anti-chicken AF488 (Invitrogen). Secondary antibody alone was used as a control. Sections were mounted with ProLong Gold antifade with DAPI (Invitrogen).

Shehata M., Waterhouse P.D., Casey A.E., Fang H., Hazelwood L, & Khokha R. (2018). Proliferative heterogeneity of murine epithelial cells in the adult mammary gland. Communications Biology, 1, 111.

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Osteoclastogenesis Assay Protocol

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Cells were cultured in αMEM containing 2 mM L-glutamine (12561), 10% heat-inactivated FBS, 100 IU/ml Penicillin and 100 IU/ml Streptomycin (Life Technologies). qPCR primers were either designed or used as previously described and tested for their distinctive product sizes (Integrated DNA Technologies) (34 (link)). For a detailed list of primer constructs see supplemental table 1. CMG-1412 media was generated as previously described (35 (link)). IF antibodies and reagents: anti-KCa3.1 (P4997) (Sigma-Aldrich), anti-CD68 (FA-11) (Biolegend) (36 (link)), Phalloidin-FITC (Sigma-Aldrich), goat anti-rabbit AF-594 and goat anti-rat AF488 (life technologies), goat serum (Gemini-Bio products). Recombinant cytokines (mM-CSF, mTNF and mRANKL) were purchased from R&D Systems; inhibitors: TRAM-34, KN-93, cyclosporine A (CsA) (Sigma-Aldrich); Fluo-4-AM (life technologies); Western blot antibodies: anti-CREB (06-863) and anti-phospho-CREB (Ser133) (06-519) (Millipore) (14 (link)), anti-CaMKIV (H-5) (Santa Cruz Technologies) (37 (link)), anti-phospho-CaMKIV (Thr196, Thr200) (PA5-37504) (Thermo Fisher Scientific), anti-NFATc1 (7A6) (Thermo Fisher Scientific) (14 (link)), anti-β-actin (13E5) (Cell Signaling Technology)

Grössinger E.M., Kang M., Bouchareychas L., Sarin R., Haudenschild D.R., Borodinsky L.N, & Adamopoulos I.E. (2017). Ca2+-dependent regulation of NFATc1 via KCa3.1 in Inflammatory Osteoclastogenesis. Journal of immunology (Baltimore, Md. : 1950), 200(2), 749-757.

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Visualizing Tumor-Infiltrating PyMT Cells

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Tumors were harvested from E-Cadherin^CFPGzmC-iCre^tdTPyMT mice and fixed in Periodate-Lysine-Paraformaldehyde (PLP) for 16–24 hours, 30% sucrose for 24 hours, then frozen in OCT. Tissue was sectioned at 20 μm thickness, blocked for 30 minutes and stained with Hoechst and a fluorescently conjugated antibody against PyMT (Thermo Fisher Scientific, catalog number MA1–46061) overnight at 4°C. Slides were washed and stained with secondary antibody (goat anti-rat AF488, Life Technologies catalog number A-11006). Images were taken on confocal microscope using 4 color channels.

Kansler E.R., Dadi S., Krishna C., Nixon B.G., Stamatiades E.G., Liu M., Kuo F., Zhang J., Zhang X., Capistrano K., Blum K.A., Weiss K., Kedl R.M., Cui G., Ikuta K., Chan T.A., Leslie C.S., Hakimi A.A, & Li M.O. (2022). Cytotoxic Innate Lymphoid Cells Sense Cancer Cell-Expressed Interleukin-15 to Suppress Human and Murine Malignancies. Nature immunology, 23(6), 904-915.

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Quantifying Immune Cell Infiltration in Tissues

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Organs were harvested and fixed in ice-cold PBS containing 4% paraformaldehyde for 18 to 24 h with gentle end-over-end mixing. Fixed organs were placed in 30% sucrose solution overnight. Saturated organs were then submerged in optimal cutting temperature compound (OCT) (Sakura) and flash-frozen in cassettes submerged in 2-methylbutane chilled with dry ice. Sections (10 μm) were permeabilized and stained with ∼1 μg/ml rabbit anti-myeloperoxidase (Abcam; no. ab9535), and 5 μg/ml rat anti-mouse CD68 (Thermo Fisher Scientific; no. 14-0681-82), followed by incubation with goat anti-rabbit AF594 (Thermo Fisher Scientific; no. A11012) and goat anti-rat AF488 (Thermo Fisher Scientific; no. A11006). Nuclei were visualized using mounting medium containing DAPI (4′,6-diamidino-2-phenylindole; Thermo Fisher Scientific). Sections were mounted on glass slides under no. 1.5 coverslips. Images were acquired with an inverted Zeiss LSM 880 confocal microscope with FAST AiryScan, using either a 10× Plan-Apochromat 0.45 NA objective or a 40× LD LCI Plan-Apochromat 1.2 NA immersion objective as indicated in the figure legend. Images were processed using the in-line AiryScan processing module in Zen Black.

Golden G.J., Toledo A.G., Marki A., Sorrentino J.T., Morris C., Riley R.J., Spliid C., Chen Q., Cornax I., Lewis N.E., Varki N., Le D., Malmström J., Karlsson C., Ley K., Nizet V, & Esko J.D. (2021). Endothelial Heparan Sulfate Mediates Hepatic Neutrophil Trafficking and Injury during Staphylococcus aureus Sepsis. mBio, 12(5), e01181-21.

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