Ly 6c apc hk1

Manufactured by Thermo Fisher Scientific

Ly-6C-APC (HK1.4) is a fluorescently labeled antibody that binds to the Ly-6C antigen. Ly-6C is a marker expressed on various cell types, including monocytes, granulocytes, and some T cell subsets. The APC (Allophycocyanin) fluorescent label allows for detection and analysis of Ly-6C-positive cells using flow cytometry.

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Lab products found in correlation

Anti f4 80 fitc bm8, by Abcam (2 mentions) Cd11b v450 m1 70, by BD (2 mentions) Facscanto 2, by BD (2 mentions) Ccr2 pe, by R&D Systems (2 mentions) Aldehyde sulfate latex beads, by Thermo Fisher Scientific (2 mentions) Collagenase dispase, by Merck Group (2 mentions) Nkg2d pe cy7 cx5, by Thermo Fisher Scientific (1 mentions) Ly6g pe 1a8, by BD (1 mentions) Facsaria 3, by BD (1 mentions) F4 80 pe cy7 bm8, by Thermo Fisher Scientific (1 mentions) Cd11c percp, by BioLegend (1 mentions) Anti cd63 h5c6, by BD (1 mentions)

3 protocols using ly 6c apc hk1

Peripheral Blood Lymphocyte Immunophenotyping

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For analysis of peripheral blood lymphocytes, tail or submandibular vein blood (< 100 μl) was collected and subjected to RBC lysis. Subsequent cell suspensions were stained with panels of antibodies: CD8α‐PE‐Cy7 (53‐6.7), CD4‐APC‐Cy7 (GK1.5), CD69‐APC (H1.2F3), CD44‐FITC (IM7), CD62L‐BV421 (MEL‐14), PD‐1‐PE (J43), NK1.1‐BV421 (PK136), FOXP3‐FITC (MF23), TNF‐α‐FITC (MP6‐XT22), and IFN‐γ–PE (XMG 1.1) (all from BD Biosciences), and NKg2D‐PE‐Cy7 (CX5) (eBioscience). For characterization of bone stromal cell populations, cells and isolation of CD11b⁺ subsets for co‐culture cells were also stained with CD11b‐BV421/BV605 (M1/70), CD3‐PE (17A2), TCR‐β–FITC (H57‐597), and Ly6G‐PE (1A8; all BD Biosciences), CD11c‐PercP (N418; Biolegend), Ly6C‐APC (HK1.4; eBioscience), and F4/80‐PeCy7 (BM8; Invitrogen). For characterization of H2‐Kb expression, RM1 cells were stained with H2‐Kb‐PE (AF6‐88.5; BD Biosciences). Data are represented as lymphocyte percentage. All analysis was performed on the FACSAria III (BD Biosciences), and data were analyzed using FlowJo 10.5.0 software (Tree star).

Owen K.L., Gearing L.J., Zanker D.J., Brockwell N.K., Khoo W.H., Roden D.L., Cmero M., Mangiola S., Hong M.K., Spurling A.J., McDonald M., Chan C., Pasam A., Lyons R.J., Duivenvoorden H.M., Ryan A., Butler L.M., Mariadason J.M., Giang Phan T., Hayes V.M., Sandhu S., Swarbrick A., Corcoran N.M., Hertzog P.J., Croucher P.I., Hovens C, & Parker B.S. (2020). Prostate cancer cell‐intrinsic interferon signaling regulates dormancy and metastatic outgrowth in bone. EMBO Reports, 21(6), e50162.

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Isolation and Characterization of Lung Macrophages

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Mouse lung tissue was digested with collagenase-dispase (sigma-Aldrich) to generate a single cell suspension as previously described ⁵⁰. Cells were then stained with the following antibodies: Anti-F4/80-FITC (BM8, Abcam), CCR2-PE (475301, R&D System), Ly-6C-APC (HK1.4, eBiosciences), CD11b-V450 (M1/70, BD Biosciences). CD11b⁺ and Ly6C^hi cells were gated within the F4/80⁺ subpopulation, and from these, CCR2⁺ cells were sorted as previously described ⁵⁰. Characterization of exosomes markers was performed as described previously ^{8 (link), 17 (link)}. Aldehyde/sulfate latex beads (Life Technology) were coupled with 30ug of exosomes, stained with FITC-conjugated anti-CD63 (H5C6, BD Pharmingen) and analyzed by FACS. Macrophages co-incubated with OFP-mitochondria labeled MSCs were incubated with anti-F4/80⁺ and double positive F4/80⁺/RFP cells sorted by FACS. Analysis was performed using a FACS Canto II (BD Biosciences) and data analyzed using FlowJo v7.6.3 software.

Phinney D.G., Di Giuseppe M., Njah J., Sala E., Shiva S., St Croix C.M., Stolz D.B., Watkins S.C., Di Y.P., Leikauf G.D., Kolls J., Riches D.W., Deiuliis G., Kaminski N., Boregowda S.V., McKenna D.H, & Ortiz L.A. (2015). Mesenchymal stem cells use extracellular vesicles to outsource mitophagy and shuttle microRNAs. Nature Communications, 6, 8472.

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Flow Cytometry Analysis of Lung Macrophages

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Mouse lung tissue was digested with collagenase-dispase (Sigma-Aldrich) to generate a single-cell suspension as previously described50 . Cells were then stained with the following antibodies: Anti-F4/80-FITC (BM8, Abcam), CCR2-PE (475301, R&D System), Ly-6C-APC (HK1.4, eBiosciences) and CD11b-V450 (M1/70, BD Biosciences). CD11b⁺ and Ly6C^hi cells were gated within the F4/80⁺ subpopulation, and from these CCR2⁺ cells were sorted as previously described50 . Characterization of exosomes markers was performed as described previously8 (link)17 (link). Aldehyde/sulfate latex beads (Life Technology) were coupled with 30 μg of exosomes, stained with fluorescein isothiocyanate -conjugated anti-CD63 (H5C6, BD Pharmingen) and analysed using FACS. Macrophages co-incubated with Orange Fluorescent Protein (OFP)-mitochondria-labelled MSCs were incubated with anti-F4/80⁺ and double-positive F4/80⁺/RFP cells sorted using FACS. Analysis was performed using a FACS Canto II (BD Biosciences) and data analysed using the FlowJo v7.6.3 software.

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