Clone ghi 61

Manufactured by BioLegend

Sourced in United States

The Clone GHI/61 is a laboratory equipment product. It is used for cell analysis and research purposes.

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Lab products found in correlation

Cd163, by BioLegend (1 mentions) Clone y1 82a, by BioLegend (1 mentions) Magnisort streptavidin positive selection beads, by Thermo Fisher Scientific (1 mentions) Clone hp6017, by BioLegend (1 mentions) Clone 19, by BD (1 mentions) Clone it2, by BD (1 mentions) 7 aad, by BD (1 mentions) Facscalibur flow cytometer, by BD (1 mentions)

3 protocols using clone ghi 61

Visualizing GM-CSFRα Expression in RA Synovial Macrophages

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To investigate GM-CSFRα expression by macrophages we incubated synovial biopsies of 5 patients with RA overnight at 4°C using primary antibodies specific for mouse antihuman GM-CSFRα (clone 4H1; Lifespan BioSciences), followed by Alexa Fluor 488 goat antimouse IgG2b antibody (Invitrogen, Carlsbad, California, USA). Subsequently, mouse monoclonal antibodies specific for macrophages (CD68; clone Y1/82A and CD163; clone GHI/61; Biolegend, San Diego, California, USA) were incubated for 60 min at room temperature and were detected using Alexa Fluor 633 goat antimouse IgG1 antibody (Invitrogen). We used a Zeiss LSM 780 Zen confocal microscope (Jena, Germany) to visualise staining.

Greven D.E., Cohen E.S., Gerlag D.M., Campbell J., Woods J., Davis N., van Nieuwenhuijze A., Lewis A., Heasmen S., McCourt M., Corkill D., Dodd A., Elvin J., Statache G., Wicks I.P., Anderson I.K., Nash A., Sleeman M.A, & Tak P.P. (2014). Preclinical characterisation of the GM-CSF receptor as a therapeutic target in rheumatoid arthritis. Annals of the Rheumatic Diseases, 74(10), 1924-1930.

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Isolation of CD163+ Exosomes from Tissues

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500 μL of magnetic beads (MagniSort Streptavidin Positive Selection Beads, Invitrogen, Catalog Number: MSPB-6003) were incubated with biotinylated anti-CD163 antibody (1:50, clone GHI/61, Biolegend), or biotinylated anti-human IgG Fc antibody (1:50, clone HP6017, Biolegend) as a control on a shaker for 1 h at room temperature. The beads were then placed on the magnet and washed three times with PBS. 1 mg of tissue-derived exosomes (in 1 mL PBS) from each sample was used in this study, of which 500 μg was collected as the control group and the other 500 μg exosomes were incubated with biotinylated anti-IgG or CD163 antibody-bounded magnetic beads overnight at 4°C. Afterward, the supernatant was transferred into a new tube for ultracentrifugation (120,000 × g for 2 h) to obtain the exosomes as the void group. The magnetic beads bound with CD163⁺ exosomes were collected, placed on the magnet, and washed 3 times with PBS to obtain CD163⁺ exosomes. The same amounts of exosomal proteins from each group were loaded for western blotting analysis. Exosomes from the control group, the void group were used for T cell suppression studies.

Altier C., Suyemoto M, & Lawhon S.D. (2000). Regulation of Salmonella enterica Serovar Typhimurium Invasion Genes by csrA. Infection and Immunity, 68(12), 6790-6797.

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Flow Cytometric Analysis of Macrophage Markers

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Macrophages from PB-purified monocytes were stained with the anti-human MHC-II conjugated to PE (clone L243, BD) or anti-human CD86 conjugated to PE (clone IT2.2; BD) or anti-human CD64 conjugated to FITC (clone 10.1; BD) or anti-human CD206 conjugated to PE (clone 19.2, BD) or anti-human DC-SIGN conjugated to FITC (clone DCN46, BD) or anti-human CD163 conjugated to PerCP (clone GHI61, Biolegend) or anti-mouse MHC-II conjugated to FITC (I-A/I-E, clone M5/114.15.2, e-Bioscience) or the corresponding isotype control antibodies and then evaluated by flow cytometry at different times post-stimulation. When possible, cells were washed and incubated with 7-AAD (BD Biosciences) for 10 minutes on ice and in the dark. The expression of the different surface markers was evaluated within the viable cell population (7-AAD negative). After labelling, the cells were analysed on a FACSCalibur flow cytometer (BD Biosciences) or Partec CyFlow (LabSystems) and the data were processed with the FlowJo 7.6.2 or vX.0.7 applications (FlowJo, LLC). Data was normalized to untreated cells, i.e., in each set of experiments, for each marker, the MFI of treatments (from the % of positive cells for each marker) was divided by MFI of media condition (from the % of positive cells for each marker).

Serafino A., Marin Franco J.L., Maio M., Trotta A., Genoula M., Castillo L.A., Birnberg Weiss F., Pittaluga J.R., Balboa L., Barrionuevo P, & Milillo M.A. (2022). Brucella abortus RNA does not polarize macrophages to a particular profile but interferes with M1 polarization. PLOS Neglected Tropical Diseases, 16(11), e0010950.

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