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Anti mouse cd45 clone 30 f11

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The Anti-mouse CD45 (clone 30-F11) is a laboratory reagent used for the detection and analysis of mouse CD45-expressing cells. It is a monoclonal antibody that binds specifically to the CD45 antigen, which is expressed on the surface of various hematopoietic cells, including lymphocytes, monocytes, and granulocytes. This reagent can be used in flow cytometry and other immunoassay applications to identify and quantify CD45-positive cells in mouse samples.

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Lab products found in correlation

Anti mouse cd4 clone gk1, by BD (3 mentions) Anti human cd45 clone hi30, by BD (3 mentions) Anti human cd3 sk7, by BioLegend (2 mentions) Anti human cd4 sk3, by BioLegend (2 mentions) Anti human cd8 sk1, by BioLegend (2 mentions) Anti mouse cd8a clone 53 6, by BD (2 mentions) Anti ki67 clone 8d5, by Cell Signaling Technology (2 mentions) Alexa fluor 568 goat anti rabbit igg h l, by Thermo Fisher Scientific (2 mentions) Alexa fluor 488 goat anti mouse igg h l, by Thermo Fisher Scientific (2 mentions) Anti human mitochondria clone 113 1, by Merck Group (2 mentions) Cd4 clone rpa t4, by BioLegend (1 mentions) Cd19 clone hib19, by BioLegend (1 mentions) Trucount, by BD (1 mentions) Anti mouse f4 80 clone bm8, by BioLegend (1 mentions) Anti mouse ifn γ xmg1, by Thermo Fisher Scientific (1 mentions) Rabbit anti ifngr1, by Merck Group (1 mentions) Anti mouse cd3 clone 17a2, by BioLegend (1 mentions) Anti mouse b220 clone ra3 6b2, by BioLegend (1 mentions) Anti mouse mhcii clone m5 114, by BioLegend (1 mentions) Anti mouse cd11b m1 70, by Thermo Fisher Scientific (1 mentions)

7 protocols using anti mouse cd45 clone 30 f11

1

Quantifying Human Leukocyte Reconstitution

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Human leukocyte reconstitution was assessed by flow cytometry using Trucount (BD Biosciences, San Jose, CA, USA) enumeration to calculate the absolute number of human B cells, CD4+ and CD8+ T cells, monocytes, NK cells, and neutrophils per μL of blood [35 (link)]. Anti-human CD45 (clone H130, BD Biosciences, San Jose, CA, USA) and anti-mouse CD45 (clone 30-F11, BD Biosciences, San Jose, CA, USA) antibodies were used to differentiate mouse from human leukocytes. Human CD45+ cells were phenotyped using antibodies specific for human CD3 (clone UCHT1, Beckman Coulter, Brea, CA, USA), CD4 (clone RPA-T4, Biolegend, San Diego, CA, USA), CD14 (clone TüK4, Invitrogen, Carlsbad, CA, USA), and CD19 (clone HIB19, Biolegend, San Diego, CA, USA). Data analysis was performed using FlowJo software (v.9.9.4, FlowJo LLC, Ashland, OR, USA).
Maidji E., Moreno M.E., Rivera J.M., Joshi P., Galkina S.A., Kosikova G., Somsouk M, & Stoddart C.A. (2019). Cellular HIV Reservoirs and Viral Rebound from the Lymphoid Compartments of 4′-Ethynyl-2-Fluoro-2′-Deoxyadenosine (EFdA)-Suppressed Humanized Mice. Viruses, 11(3), 256.
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2

Immunoblotting and Flow Cytometry Protocols

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For immunoblotting, these antibodies are used: rabbit anti-STAT1 (CST, #14994), rabbit anti–phospho-STAT1 (CST, #7649), rabbit anti-EZH2 (CST, #5246), mouse anti–MHC-I class I (Santa Cruz Biotechnology, #sc-55582), rabbit anti-IFNGR1 (Millipore, #MABF753), and mouse anti–β-actin (Sigma-Aldrich, #A5441).
For flow cytometry, the following fluorochrome-conjugated antibodies are used: anti-mouse H-2Kb/H-2Db (clone 28-8-6, BioLegend), anti-mouse H-2Kb bound to SIINFEKL (clone 25-D1.16, BioLegend), anti-mouse CD3 (clone 17A2, BioLegend), anti-mouse CD28 (clone 37.51, BioLegend), anti-mouse granzyme B (clone GB11, BioLegend), anti-mouse B220 (clone RA3-6B2, BioLegend), anti-mouse CD49b (clone DX5, BioLegend), anti-mouse Gr-1 (clone RB6-8C5, BioLegend), anti-mouse MHC-II (clone M5/114.15.2, BioLegend), anti-mouse F4/80 (clone BM8, BioLegend), anti-mouse IFN-γ (XMG1.2, eBioscience), anti-mouse CD11b (M1/70, eBioscience), anti-mouse NK1.1 (clone PK136, BD), anti-mouse CD103 (clone M290, BD), anti-mouse CD206 (clone MR5D3, BD), anti-mouse CD24 (clone M1/69, BD), anti-mouse CD4 (clone GK1.5, BD), anti-mouse CD45 (clone 30-F11, BD), anti-mouse CD8a (clone 53-6.7, BD), anti-mouse Foxp3 (clone MF23, BD), anti-mouse γδ TCR (clone GL3, BD), anti-human CD3 (SK7, BioLegend), anti-human CD4 (SK3, BioLegend), anti-human CD8 (SK1, BioLegend), and anti-human 137(4-1BB) (4B4-1, BioLegend).
Guo W., Wang Y., Yang M., Wang Z., Wang Y., Chaurasia S., Wu Z., Zhang M., Yadav G.S., Rathod S., Concha-Benavente F., Fernandez C., Li S., Xie W., Ferris R.L., Kammula U.S., Lu B, & Yang D. (2021). LincRNA-immunity landscape analysis identifies EPIC1 as a regulator of tumor immune evasion and immunotherapy resistance. Science Advances, 7(7), eabb3555.
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3

Evaluating Tumor-Infiltrating Immune Cells

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Yumm cells (3.105) were intradermally injected into C57BL/6 mice. At day 11, mice were sacrificed and tumors were collected and digested with Mouse Tumor Dissociation kit and GentleMacs (Miltenyi). Cells were counted and directly stained with LIVE/DEAD reactive dyes (Invitrogen, 1:100) and the indicated antibodies: anti-mouse CD45 (clone 30-F11, 1:200), anti-mouse CD8 (clone 53-6.7, 1:200), anti-mouse CD4 (clone GK1.5, 1/200), anti-mouse Ki67 (clone B56, 1:5) and anti-mouse IFN-γ (clone XMG1.2, 1:100) are from BD Biosciences; anti-mouse Foxp3 (clone FJK-16s, 1:50), anti-mouse CTLA-4 (clone UC10-4B9, 1:200) and anti-mouse PD-1 (clone J43, 1:200) are from Thermo Fisher; anti-mouse TIM-3 (clone 1G9, 1:200), anti-mouse TIGIT (clone 1G9, 1:200), anti-mouse CD226 (clone TX42.1, 1:200) and anti-mouse Thy1 (clone 30-H12, 1:400) are from Biolegend. Cell suspensions were incubated with anti-CD16/CD32 blocking antibodies (Thermo Fisher) prior to incubation. For IFN-γ analysis, tumor cells were incubated with a Cell Stimulation Cocktail (Invitrogen). Cells were analyzed with a BD LSR Fortessa X-20 (BD Biosciences), followed by Flow-Jo software analysis. Gating strategies followed for flow cytometry analysis are shown in Supplementary Fig. 1.
Imbert C., Montfort A., Fraisse M., Marcheteau E., Gilhodes J., Martin E., Bertrand F., Marcellin M., Burlet-Schiltz O., Peredo A.G., Garcia V., Carpentier S., Tartare-Deckert S., Brousset P., Rochaix P., Puisset F., Filleron T., Meyer N., Lamant L., Levade T., Ségui B., Andrieu-Abadie N, & Colacios C. (2020). Resistance of melanoma to immune checkpoint inhibitors is overcome by targeting the sphingosine kinase-1. Nature Communications, 11, 437.
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4

Mouse Tumor Cell and PBMC Culture

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Mouse Luise Lung carcinoma (LLC) and B16 cell line were a kind gift from B. Lu’s laboratory, were cultured in DMEM high glucose (Hyclone) with 10 percent heal inactivated FBS (Gibco), 2 mM L-glutamine (Invitrogen, Carlsbad, CA), 50 U/penicillin (Invitrogen), streptomycin (50 μg/ml) (Invitrogen). Peripheral blood mononuclear cells (PBMCs) transduced with TCR gp100 and 526-MEL were obtained from Dr. U. Kammula’s laboratory. PBMCs were cultured in complete medium [RPMI-1640, 10% heat-inactivated human AB serum (Gemini Bio-Products, Woodland, CA), 2 mM L-glutamine (Invitrogen, Carlsbad, CA), 50 U/penicillin (Invitrogen), streptomycin (50 μg/ml) (Invitrogen), gentamicin (50 μg/ml) (Invitrogen), 10 mM HEPESs (Invitrogen), and Amphotericin B (250 ng/ml) (Invitrogen). Six-week female C57BL/6j mice and nude mice were purchased from Jackson laboratory. anti-mouse CD45 (clone 30-F11, BD), anti-mouse CD8a (clone 53-6.7, BD), anti-mouse CD4 (clone GK1.5, BD) CD69 (clone H1.2F3, Biolegend), anti-human CD3 (SK7, BioLegend), anti-human CD4 (SK3, BioLegend), anti-human CD8 (SK1, BioLegend), and anti-human 137(4-1BB) (4B4-1, BioLegend).
Bhuniya A., Pattarayan D, & Yang D. (2022). Lentiviral vector transduction provides a nonspecific immunogenicity for syngeneic tumor models. Molecular carcinogenesis, 61(12), 1073-1081.
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5

Flow Cytometric Characterization of CAR T-Cells

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Flow cytometry was performed on BD LSR Fortessa. Transduction efficiency was measured with an APC-conjugated antibody toward NGFR (CD271; clone ME20.4, BioLegend) and Protein L for CCR, whereas for CARs it was measured in the phycoerythrin (PE) channel to detect dsRed and AffiniPure F(ab’)2 Fragment Goat Anti-Mouse IgG (Jackson ImmunoResearch). The following antibodies were used for flow cytometry staining: Anti-Human CD3 (clone UCHT1, BD Biosciences), Anti-Human CD4 (clone L200, BD Biosciences), Anti-Mouse CD45 (clone 30-F11, BD Biosciences), Anti-Human CD8 (clone SK1, BD Biosciences), Anti-Human CD45 (clone HI30, BD Biosciences), Anti-human CD366 (TIM-3) (clone F38-2E2, BioLegend or clone F38-2E2, Thermo Fisher Scientific), Anti-human CD279 (PD-1) (clone EH12.2H7, BioLegend or clone eBioJ105, Thermo Fisher Scientific), Anti-human CD62L (clone DREG-56, BioLegend), Anti-human CD269 (BCMA) (clone 19F2, BioLegend), Anti-human CD38 (clone HIT2, BioLegend), CD45RA (clone HI100, Thermo Fisher Scientific), LAG-3 (CD223; clone 3DS223H, Thermo Fisher Scientific) (staining 1 μg per mL of sample, incubation for 30 minutes at 4°C in PBS with 1% BSA, 10% FBS and 0.1% NaN3 sodium azide). For the quantification of the CD38, the Quantibrite Phycoerythrin (PE) Fluorescence Quantitation Kit (BD) was used. Flow cytometry data analysis was performed with FCS Express 6 flow software.
Katsarou A., Sjöstrand M., Naik J., Mansilla-Soto J., Kefala D., Kladis G., Nianias A., Ruiter R., Poels R., Sarkar I., Patankar Y.R., Merino E., Reijmers R.M., Frerichs K.A., Yuan H., de Bruijn J., Stroopinsky D., Avigan D., van de Donk N.W., Zweegman S., Mutis T., Sadelain M., Groen R.W, & Themeli M. (2021). Combining a CAR and a chimeric costimulatory receptor enhances T cell sensitivity to low antigen density and promotes persistence. Science translational medicine, 13(623), eabh1962.
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6

Immunofluorescence Staining of Cells

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Cells were fixed, permeabilized, blocked against nonspecific adhesion, immunostained for target proteins, and then imaged on an inverted Eclipse Ti epifluorescence microscope (Nikon). Primary antibodies were administered at manufacturer recommended concentration: anti-human CD45 (clone HI30, Becton Dickinson), anti-mouse CD45 (clone 30-F11, Becton Dickinson), anti-human mitochondria (clone 113–1, MilliporeSigma), anti-Ki67 (clone 8D5, Cell Signaling Technology), and anti-phospho-histone H2A.X (Ser139, clone 20E3, Cell Signaling Technology). The secondary antibodies used were Alexa Fluor 488 goat anti-mouse IgG (H+L) (Invitrogen) and Alexa Fluor 568 goat anti-rabbit IgG (H+L) (Invitrogen).
Yankaskas C.L., Thompson K.N., Paul C.D., Vitolo M.I., Mistriotis P., Mahendra A., Bajpai V.K., Shea D.J., Manto K.M., Chai A.C., Varadarajan N., Kontrogianni-Konstantopoulos A., Martin S.S, & Konstantopoulos K. (2019). A microfluidic assay for the quantification of the metastatic propensity of breast-cancer specimens. Nature biomedical engineering, 3(6), 452-465.
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7

Immunofluorescence Staining of Cells

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Cells were fixed, permeabilized, blocked against nonspecific adhesion, immunostained for target proteins, and then imaged on an inverted Eclipse Ti epifluorescence microscope (Nikon). Primary antibodies were administered at manufacturer recommended concentration: anti-human CD45 (clone HI30, Becton Dickinson), anti-mouse CD45 (clone 30-F11, Becton Dickinson), anti-human mitochondria (clone 113–1, MilliporeSigma), anti-Ki67 (clone 8D5, Cell Signaling Technology), and anti-phospho-histone H2A.X (Ser139, clone 20E3, Cell Signaling Technology). The secondary antibodies used were Alexa Fluor 488 goat anti-mouse IgG (H+L) (Invitrogen) and Alexa Fluor 568 goat anti-rabbit IgG (H+L) (Invitrogen).
Yankaskas C.L., Thompson K.N., Paul C.D., Vitolo M.I., Mistriotis P., Mahendra A., Bajpai V.K., Shea D.J., Manto K.M., Chai A.C., Varadarajan N., Kontrogianni-Konstantopoulos A., Martin S.S, & Konstantopoulos K. (2019). A microfluidic assay for the quantification of the metastatic propensity of breast-cancer specimens. Nature biomedical engineering, 3(6), 452-465.
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