F4 80 bm8

Manufactured by Thermo Fisher Scientific

Sourced in United States

The F4/80 (BM8) is a cell surface glycoprotein that is expressed on mature tissue macrophages. It is commonly used as a marker for the identification and analysis of macrophages in various research applications.

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Lab products found in correlation

Cd11b m1 70, by Thermo Fisher Scientific (13 mentions) Siglecf e50 2440, by BD (8 mentions) Facscanto, by BD (7 mentions) Facsdiva software, by BD (6 mentions) Cd11c n418, by BioLegend (6 mentions) Cd11c n418, by Thermo Fisher Scientific (6 mentions) Ly6g 1a8, by BioLegend (6 mentions) Cd11c hl3, by BD (5 mentions) Ly6c hk1, by BioLegend (5 mentions) Cd8 53 6, by BioLegend (5 mentions) Cd11b m1 70, by BD (5 mentions) Facscanto 2, by BD (5 mentions) Foxp3 fjk 16s, by Thermo Fisher Scientific (4 mentions) Antigen unmasking solution, by Vector Laboratories (4 mentions) Cd11b, by BD (4 mentions) Cd64 x54 5 7, by BioLegend (4 mentions) Ly6c al 21, by BD (4 mentions) Lsrii flow cytometer, by BD (4 mentions) Lsm 710, by Zeiss (4 mentions) Cd8 53 6, by Thermo Fisher Scientific (3 mentions)

74 protocols using f4 80 bm8

Immunohistochemical Analysis of Spleen

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Splenic tissue from WT and Dock8^−/− mice were fixed in 4% paraformaldehyde overnight at
4° C. After washing in PBS, tissues were dehydrated through sequential exposure to solutions of 10%, 20% and 30%
sucrose, mounted in acryomold with O.C.T. Compound (Tissue-Tek, Sakura), and stored at −80°C prior to sectioning
(7 μm) and staining. The following antibodies were used for staining different cell subsets: CD11c (N418), CD169
(MOMA), TCRβ (H57–597), F4/80 (BM8, Invitrogen), Fibroblast Marker (ER-TR7, Santa Cruz Biotechnology), SIGN-R1
(ER-TR9), B220 (RA3–6B2). The images were acquired immediately after stain with the Nikon eclipse Ti microscope using
20x objectives.

Liu D., Yin X., Olyha S.J., Nascimento M.S., Chen P., White T., Gowthaman U., Zhang T., Gertie J.A., Zhang B., Xu L., Yurieva M., Devine L., Williams A, & Eisenbarth S.C. (2019). IL-10-Dependent Crosstalk between Murine Marginal Zone B Cells, Macrophages, and CD8α+ Dendritic Cells Promotes Listeria monocytogenes Infection. Immunity, 51(1), 64-76.e7.

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Immunofluorescence Staining of Frozen Tissue Sections

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For immunofluorescence staining, primary antibodies were diluted in 1% PBS-BSA. Frozen sections were incubated with primary antibodies for Agrin (R&D Systems, #AF550), Hu (abcam, #96474), F4/80(BM8) (Invitrogen, #41-4801-82), Col4 (abcam, #236640), Atf4 (Invitrogen, #MA5-32364), Bax1 (SCBT, #sc-7480), Egr1 (Invitrogen, #MA5-15008), IL-18 (Invitrogen, #MA5-47203) and Hif1a (Invitrogen, #PA116601) overnight at 4 °C, followed by secondary antibodies (Alexa Fluor 647 conjugated anti-rabbit IgG; Alexa Fluor 555 conjugated anti-goat IgG; Alexa fluor 488 conjugated anti-rat; Invitrogen) for 1 h. Cell nuclei were visualized by DAPI. Sections were covered with aqueous Poly/Mount (Polyscience Inc.) and examined with a Zeiss LSM 780 laser-scanning confocal microscope (Zeiss). Images were compiled by ImageJ and Adobe Photoshop CS6 software package.

Ferenczi S., Mogor F., Takacs P., Kovacs T., Toth V.E., Varga Z.V., Kovács K., Lohinai Z., Vass K.C., Nagy N, & Dora D. (2023). Depletion of muscularis macrophages ameliorates inflammation-driven dysmotility in murine colitis model. Scientific Reports, 13, 22451.

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Immunofluorescence Staining of Frozen Tissue Sections

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For immunofluorescence staining, primary antibodies were diluted in 1 % PBS-bovine serum albumin. Frozen sections were incubated with primary antibodies for agrin (R&D Systems, Minneapolis, MN; #AF550), Hu (Abcam, Cambridge, MA; #96474), CD45 (BioLegend, San Diego, CA; #30F11), anti-GFP (R&D Systems; #AF4240), CD11b (Bio-Rad, Hercules, CA; #M1/70.15), CSF1R (Invitrogen, Waltham, MA; #PA5-25974), F4/80(BM8) (Invitrogen; #41-4801-82), Iba1 (Invitrogen; #PA5-27436), NCAM (Invitrogen; #PA5-78402), Col4 (Abcam; #236640) and S100A1 (Invitrogen; #PA1-932), Ly6G (Abcam; #25377), MPO (R&D Systems; #AF3667), CD45 (BioLegend; #103102), G-SMA (SBC; #65638), TNF (Abcam; #183218), MMP2 (Invitrogen; #PA5-115583), and MMP10 (Invitrogen; #PA5-79677) overnight at 4°C, followed by secondary antibodies (Alexa Fluor 647, 546, and 488 conjugated anti-rabbit immunoglobulin G; Alexa Fluor 647 and 555 conjugated anti-goat immunoglobulin G; Alexa Fluor 488 and 594 conjugated anti-rat; Invitrogen) for 1 hour. Cell nuclei were visualized by DAPI. Sections were covered with aqueous Poly/Mount (Polyscience Inc) and examined with a Zeiss LSM 780 laser-scanning confocal microscope (Zeiss, Oberkochen) or Nikon Eclipse Ti2-E inverted microscope with an Abberior STED super resolution imaging platform. Images were compiled by ImageJ and Adobe Photoshop CS6 (San Diego, CA) software package.

Dora D., Ferenczi S., Stavely R., Toth V.E., Varga Z.V., Kovacs T., Bodi I., Hotta R., Kovacs K.J., Goldstein A.M, & Nagy N. (2021). Evidence of a Myenteric Plexus Barrier and Its Macrophage-Dependent Degradation During Murine Colitis: Implications in Enteric Neuroinflammation. Cellular and Molecular Gastroenterology and Hepatology, 12(5), 1617-1641.

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Tumor and Immune Cell Analysis Protocol

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For tumor KIT⁺ cell and macrophage analysis, tumors were minced, incubated in 12 mg/mL type 2 collagenase (Worthington Biochemical) plus 0.5 mg/mL DNAse I (Roche Diagnostics) for 30 minutes at 37°C, then washed through 100 then 40 μm nylon cell strainers (Falcon, BD Biosciences) with 1% fetal calf serum (FCS). For T cell analysis, tumors and draining lymph nodes (DLN) were mechanically dissociated as described (14 (link)). Spleens were mashed through a 70 μm strainer, incubated in ammonium chloride lysis buffer (eBioscience), quenched in 1% FCS, then washed through a 40 μm strainer. Bone marrow was flushed from one tibia per mouse, pooled by treatment group, homogenized by repeated aspiration through an 18-gauge needle, and then washed in 1% FCS. All cells were analyzed on a FACSAria (BD Biosciences) as described (14 (link)). Mouse-specific antibodies included CD45 (clone 30-F11), Kit (CD117) (2B8), CD11b (M1/70), CD3 (145-2C11), CD4 (GK1.5), Ly6C (AL-21), and Ly6G (1A8) from BD Biosciences; F4/80 (BM8) from Invitrogen; CD45 (30-F11), Kit (Ack2), CD8 (53–6.7), and FoxP3 (FJK-16s) from eBioscience. Intracellular staining for FoxP3 was performed using the FoxP3 Staining Buffer Set (eBioscience). Human-specific KIT antibody (YB5.B8) was purchased from BD Biosciences.

Kim T.S., Cavnar M.J., Cohen N.A., Sorenson E.C., Greer J.B., Seifert A.M., Crawley M.H., Green B.L., Popow R., Pillarsetty N., Veach D.R., Ku A.T., Rossi F., Besmer P., Antonescu C.R., Zeng S, & DeMatteo R.P. (2014). Increased KIT inhibition enhances therapeutic efficacy in gastrointestinal stromal tumor. Clinical cancer research : an official journal of the American Association for Cancer Research, 20(9), 2350-2362.

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Comprehensive T Cell Immunophenotyping

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T cells were interrogated with allophycocyanin (APC)–conjugated anti-CD25 (PC61.5), eFluor 450-conjugated anti-CD73 (eBioTY/11.8), PE-Cy7–conjugated anti-FR4 (eBio12A5), Alexa Fluor 700–conjugated anti-CD44 (IM7), PerCP-Cy5.5-conjugated anti-CD45.1 (A20), V500–conjugated anti-CD4 (RM4-5) (eBiosciences). T cells suspensions were also stained with APC–eFluor780–conjugated anti-B220 (RA3-6B2), -CD11b (MI-70), -CD11c (N418) (all eBiosciences), and -F4/80 (BM8; Invitrogen, Carlsbad, CA), for use as dump channel reagents. Stained T cells were then treated with Foxp3 Fixation/Permeabilization Concentrate and Diluent and stained with APC–conjugated anti-Foxp3 (FJK-16s) (eBioscience). In some experiments cells were also stained with Pacific Blue–conjugated T-bet (BioLegend, San Diego, CA) following fixation/permeabilization.

Kalekar L.A., Schmiel S.E., Nandiwada S.L., Lam W.Y., Barsness L.O., Zhang N., Stritesky G.L., Malhotra D., Pauken K.E., Linehan J.L., O’Sullivan M.G., Fife B.T., Hogquist K.A., Jenkins M.K, & Mueller D.L. (2016). CD4+ T cell anergy prevents autoimmunity and generates regulatory T cell precursors. Nature immunology, 17(3), 304-314.

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Comprehensive T Cell Immunophenotyping

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Multicolor Flow Cytometry Analysis of Murine Splenocytes

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Spleen single-cell suspensions were incubated with a Fc-receptor blocking antibody 2.4G2 (kindly provided by Dr. Louis Boon, Bioceros BV, Utrecht, The Netherlands) and the percentage of spleen cell populations (macrophages, B cells, Dendritic cells, T cells, monocytes) were determined using the following antibodies: F4/80 (BM8) (Invitrogen, Bleiswijk, The Netherlands), CD45R (B220) (eBioscience, Vienna, Austria), CD11c (HL3) (BD Biosciences), CD4 (GK1.5) (eBioscience, Vienna, Austria), CD8α (53-6.7) (eBioscience, Vienna, Austria), CD3e (145-2C11) (eBioscience, Vienna, Austria), Lineage (Lin) (B220, NK1.1, CD90, CD49. Ly6G) (ebioscience, Vienna, Austria) CD45.1 (ebioscience, Vienna, Austria). Samples were analyzed with a LSR Fortessa II (Beckman Coulter) and the FlowJo software (Tree Star Inc., Ashland, The United States). The different spleen cell populations were defined as follow: DCs (CD11c+MHCII+), macrophages (F4/80+), B cells (CD45R+MHCII+), CD4+ (CD3+CD8-CD4+), CD8+T cells (CD3+CD8+CD4-) and monocytes (CD45+Lin-F4/80-CD11c-MHCII-CD11b+Ly-6C+). Percentages were reported to the total number of splenocytes of each mouse to calculate the number of cells per population (Figure S2 and S3).

Costes L.M., van der Vliet J., Farro G., Matteoli G., van Bree S.H., Olivier B.J., Nolte M.A., Boeckxstaens G.E, & Cailotto C. (2014). The Spleen Responds to Intestinal Manipulation but Does Not Participate in the Inflammatory Response in a Mouse Model of Postoperative Ileus. PLoS ONE, 9(7), e102211.

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Multiparameter Immune Cell Analysis

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Single-cell suspensions were prepared (Tumor Dissociation Kit, 130-096-730, Miltenyi Biotec, Paris, France) and incubated with fluorophore-conjugated anti-mouse antibodies (CD45 [30-F11], CD3 [17-A2], CD4 [RM4-5], CD8 [53-6.7], CD11b [M1/70], Ly-6g [1A8]; all from BD Biosciences, Le Pont de Claix, France), CD206 (C068C2) (from BioLegend EU Amsterdam, NL), and F4/80 (BM8; from ThermoFisher Scientific). Data acquisition was performed on the Fortessa LSR (BD Biosciences) and BD FacsDiva software was used for analysis, as published previously.⁴⁷

Samain R., Brunel A., Douché T., Fanjul M., Cassant-Sourdy S., Rochotte J., Cros J., Neuzillet C., Raffenne J., Duluc C., Perraud A., Nigri J., Gigoux V., Bieche I., Ponzo M., Carpentier G., Cascone I., Tomasini R., Schmid H.A., Mathonnet M., Nicolle R., Bousquet M.P., Martineau Y., Pyronnet S., Jean C, & Bousquet C. (2021). Pharmacologic Normalization of Pancreatic Cancer-Associated Fibroblast Secretome Impairs Prometastatic Cross-Talk With Macrophages. Cellular and Molecular Gastroenterology and Hepatology, 11(5), 1405-1436.

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Isolation and Staining of Liver Non-Parenchymal Cells

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Liver non-parenchymal cells were isolated and stained as previously described [34 (link)]. Cells were stained with fluorophore-conjugated antibodies directed against the following surface antigens: CD45 (30-F11; Biolegend), Ly6G (1A8; ThermoFisher (eBioscience)), CD11b (M1/70; ThermoFisher (eBioscience)) and F4/80 (BM8; ThermoFisher (eBioscience)). Flow cytometry was performed using a BD LSRII instrument and data were analyzed with Flow Jo V10 software.

McMahan R.H., Najarro K.M., Mullen J.E., Paul M.T., Orlicky D.J., Hulsebus H.J, & Kovacs E.J. (2021). A novel murine model of multi-day moderate ethanol exposure reveals increased intestinal dysfunction and liver inflammation with age. Immunity & Ageing : I & A, 18, 37.

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Isolation and Analysis of Kidney Macrophages

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At sacrifice 10 days after IRI, kidneys were harvested from Lyz-met+/+ and Lyz-met−/− mice following perfusion with PBS. Kidneys were cut into small pieces and digested at 37°C in 1 mg/ml collagenase IV (Worthington) in complete RPMI for 30 minutes. Cells were filtered through 70 mm cell strainers and washed twice in PBS. The single-cell suspensions were slowly layered over 5 mL of Lymphocyte (Cedarlane). The tubes were spun for 1,300xg for 30 minutes at room temperature. The cell layer at the interface of the media and Lymphocyte was collected and washed twice with PBS and used for FACS staining. To identify the macrophage population, cells were stained with fluorescent-conjugated antibodies CD11b (M1/70, #14-0112-82) and F4/80 (BM8, #14-4801-82, eBioscience^TM, ThermoFisher Scientific, Waltham, MA). Data were acquired on a BD LSRFortessa cytometer (BD Biosciences) and analyzed with FlowJo (Tree Star Inc.), as described previously (Ramani et al., 2018 ).

Fu H., Gui Y., Liu S., Wang Y., Bastacky S.I., Qiao Y., Zhang R., Bonin C., Hargis G., Yu Y., Kreutzer D.L., Biswas P.S., Zhou Y., Wang Y., Tian X.J., Liu Y, & Zhou D. (2021). The hepatocyte growth factor/c-met pathway is a key determinant of the fibrotic kidney local microenvironment. iScience, 24(10), 103112.

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