Anti cd3e clone 145 2c11

Manufactured by BioLegend

The Anti-CD3e (clone 145-2C11) is a lab equipment product that specifically binds to the CD3e subunit of the T cell receptor complex. It is a widely used tool in immunology research to activate and study T cell responses.

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Lab products found in correlation

Anti cd11b clone m1 70, by BioLegend (5 mentions) Anti cd45.2 clone 104, by BioLegend (4 mentions) Anti cd4 clone gk1.5, by BD (3 mentions) Anti cd8 clone 53 6, by BD (3 mentions) Calibrite apc beads, by BD (3 mentions) Anti ter119 clone ter 119, by BioLegend (3 mentions) Anti cd27 clone lg 3a10, by BioLegend (2 mentions) Anti cd19 clone 1d3, by BD (2 mentions) Anti cd4 clone rm4 5, by Thermo Fisher Scientific (2 mentions) Anti cd127 clone b12 1, by BD (2 mentions) Pe cy7 conjugated anti sca 1 clone d7, by BD (2 mentions) Brilliant violet 785 conjugated anti ly6c clone hk1, by BioLegend (2 mentions) Percp cy5.5 conjugated anti ly6g clone 1a8, by BioLegend (2 mentions) Brilliant violet 421 conjugated anti cd34 clone ram34, by BD (2 mentions) Apc conjugated anti cd117 clone 2b8, by BioLegend (2 mentions) Brilliant violet 605 conjugated anti cd115 clone t38 320, by BD (2 mentions) Anti b220 clone ra3 6b2, by BioLegend (2 mentions) Anti cd19 clone 6d5, by BioLegend (2 mentions) Anti cd8 clone 53 6, by BioLegend (2 mentions) Anti cd11b clone m1 70, by BD (2 mentions)

9 protocols using anti cd3e clone 145 2c11

Quantification of Spinal Cord Immune Cells

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Single-cell suspensions from spinal cords were obtained via mechanical dissociation on a cell strainer. Immune cells were separated over a two-phase Percoll-density gradient. Staining of αβTCR/CD4⁺ T cells, αβTCR/CD8⁺ T cells and CD45/CD11b cells (macrophages/microglia) was performed using the following antibodies in a 1:200 dilution: Anti-CD3e (clone 145-2C11), BioLegend; anti-CD4 (clone GK 1.5), BD; anti-CD8 (clone 53-6.7), BD; anti-CD8 (clone 53–6.7), BD; anti-CD11b (clone M1/70), BioLegend; anti-CD45.2 (clone 104), BioLegend. The addition of Calibrite APC beads (BD) allowed for cell quantification. Flow cytometry was performed using a FACSCalibur operated by Cell Quest software (Becton Dickinson).

Berghoff S.A., Gerndt N., Winchenbach J., Stumpf S.K., Hosang L., Odoardi F., Ruhwedel T., Böhler C., Barrette B., Stassart R., Liebetanz D., Dibaj P., Möbius W., Edgar J.M, & Saher G. (2017). Dietary cholesterol promotes repair of demyelinated lesions in the adult brain. Nature Communications, 8, 14241.

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Comprehensive Immune Cell Profiling by Flow Cytometry

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Flow cytometry was performed as previously described Fang et al. (2010 (link)). The following antibodies were used: anti-CD3e (clone 145-2C11, Biolegend), anti-CD11b (clone M1/70, Biolegend), anti-CD19 (clone 1D3, BD Biosciences), anti-CD27 (clone LG.3A10, Biolegend), anti-CD29 (clone HMβ1-1, Biolegend San Diego, CA), anti-CD44 (clone IM7, Biolegend), anti-CD45.1 (clone A20, Biolegend), anti-CD45.2 (clone 104, Biolegend), anti-CD49a (clone HMα1, Biolegend), anti-CD49b (clone DX5, Biolegend), anti-CD105 (clone MJ7/18, Biolegend), anti-CD106 (clone 429, Biolegend), anti-BrDU (clone PRB-1, eBioscience), anti-CXCR3 (clone CXCR3-173, Biolegend), anti-Eomes (clone Dan11mag, eBioscience), anti-KLRG1 (clone 2F1/KLRG1, Biolegend), anti-Ly-6A/E (Sca-1) (clone E13-161.7, Biolegend), anti-Ly49A (clone A1/Ly49A, Biolegend), anti-Ly49C/I (clone 5E6, Biolegend), anti-Ly49D (clone 4E5, Biolegend), anti-Ly49G2 (clone LGL-1, eBioscience), anti-Ly49H (clone 3D10, Biolegend), anti-NK1.1 (clone PK136, Biolegend), NKG2A/C/E (clone 20d5, Biolegend), anti-Tbet (clone 4B10, Biolegend), and anti-TRAIL (clone N2B2, Biolegend). At least 500 000 cells were analyzed by flow cytometry at the Fox Chase Cell Sorting Facility using an LSR II system (BD Biosciences).

Nair S., Fang M, & Sigal L.J. (2014). The natural killer cell dysfunction of aged mice is due to the bone marrow stroma and is not restored by IL-15/IL-15Rα treatment. Aging Cell, 14(2), 180-190.

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Immunophenotypic Analysis of Murine Hematopoietic Stem Cells

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Miltenyi AutoMacs CD117-enriched bone marrow cells were stained for 1 h at 20°C protected from light with a mix of biotin-conjugated antibodies (including anti-CD3e (clone 145–2C11, BioLegend), anti-CD4 (clone RM4–5, Thermo Fisher), anti-CD8 (clone 53–6.7), anti-CD19 (clone 6.D5, BioLegend), anti-CD127 (clone B12–1, BD), anti-B220 (clone RA3–6B2, BioLegend), and anti-Ter119 (clone TER-119, BioLegend)), as well as PE-Cy7-conjugated anti-SCA-1 (clone D7, BD), Brilliant Violet 785-conjugated anti-LY6C (clone HK1.4, BioLegend), PerCp Cy5.5-conjugated anti-LY6G (clone 1A8, BioLegend), Brilliant Violet 421-conjugated anti-CD34 (clone RAM34, BD), APC-conjugated anti-CD117 (clone 2B8, BioLegend), and Brilliant Violet 605-conjugated anti-CD115 (clone T38–320, BD), PE- Cy5 conjugated CD135 and PerCP-eFluor710-conjugated anti-CD16/CD32 (clone 93, Thermo Fisher). Cells were then washed twice with FACS buffer and incubated for 15 min at 20°C with streptavidin-APC-Cy7 (BD). Cells were washed once in FACS buffer, and resuspended in FACS buffer for analyses.

Schnabel E.L, & Jones A.L. (2001). Isolation and Characterization of Five Erwinia amylovora Bacteriophages and Assessment of Phage Resistance in Strains of Erwinia amylovora. Applied and Environmental Microbiology, 67(1), 59-64.

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Isolation and Sorting of Myeloid Progenitor Cells

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AutoMacs CD117-enriched BM cells were stained for 1 h at 20°C protected from light with a mix of biotin-conjugated antibodies (including anti-CD3e (clone 145–2C11, BioLegend), anti-CD4 (clone RM4–5, Thermo Fisher), anti-CD8 (clone 53–6.7), anti-CD19 (clone 6.D5, BioLegend), anti-CD127 (clone B12–1, BD), anti-B220 (clone RA3–6B2, BioLegend), and anti-Ter119 (clone TER-119, BioLegend)), together with PE-Cy7-conjugated anti-SCA-1 (clone D7, BD), Brilliant Violet 785-conjugated anti-LY6C (clone HK1.4, BioLegend), PerCp Cy5.5-conjugated anti-LY6G (clone 1A8, BioLegend), Brilliant Violet 421-conjugated anti-CD34 (clone RAM34, BD), APC-conjugated anti-CD117 (clone 2B8, BioLegend) and Brilliant Violet 605-conjugated anti-CD115 (clone T38–320, BD), PE- Cy5 conjugated CD135 and BUV395-conjugated anti-CD16/CD32 (clone 93, Thermo Fisher). MDPs (Lin-, KIT+, SCA-, CD16/32-, CD34hi, CD135+, CD115+ ), GMP(Lin-, KIT+, SCA-, CD16/32+,CD34hi) and C-GMP (Lin-, KIT+, SCA-, CD16/32+/−,CD34hi) (Fig. 3A), were sorted using FACSAria II (Becton, Dickenson, and Company) and cells were collected in a solution of DPBS + 50% FBS (Atlanta Biologicals). Other granulocytic and monocytic lineage progenitors stages were sorted based on the gating strategy shown in Extended data Fig.1E.

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Comprehensive Immune Cell Profiling by Flow Cytometry

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Peritoneal exudate cells were harvested by peritoneal lavage with 10 ml ice cold FACS buffer (PBS supplemented with 2% FBS and 50 U/ml Penicillin and 50 µg/ml Streptomycin). Splenocytes were isolated by filtering through two 100 μm cell strainers into 10 ml ice cold FACS buffer. Residual red blood cells were lysed with Red Blood Cell lysis buffer (Sigma, St Louis, MO), counted and stained for flow cytometry.
Cells were incubated with FACS buffer plus 1% rat serum, 1% hamster serum and 1% Fc-block for 15 min. Surface staining was performed for 30 min at room temperature. Cells were then washed and fixed with 2% formaldehyde. Cells were analyzed on an LSRII or LSR Fortessa flow cytometer (BD, Franklin Lakes, NJ) and the data were analyzed using FlowJo software (Tree Star, Inc, Ashland, OR). The following antibodies were used: anti-CD3e (clone 145-2C11, Biolegend, San Diego, CA), anti-CD4 (clone RM4-5, BD Pharmingen), anti-CD8 (clone 53-6.7, Biolegend), anti-IgM (clone II/41, BD Pharmingen), anti-CD19 (clone 6D5, Biolegend), anti-NK1.1 (clone PK136, BD Pharmingen), anti-NKp46 (clone 29A1.4, eBioscience, San Diego, CA), anti-Ly6C (clone HK1.4, Biolegend), anti-Ly6G (clone 1A8, Biolgend, anti-CD11b (clone M1/70, BD Pharmingen), anti-F4/80 (clone BM8, Biolegend).

MacDuff D.A., Reese T.A., Kimmey J.M., Weiss L.A., Song C., Zhang X., Kambal A., Duan E., Carrero J.A., Boisson B., Laplantine E., Israel A., Picard C., Colonna M., Edelson B.T., Sibley L.D., Stallings C.L., Casanova J.L., Iwai K, & Virgin H.W. (2015). Phenotypic complementation of genetic immunodeficiency by chronic herpesvirus infection. eLife, 4, e04494.

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Detailed FACS Sorting and Analysis

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For FACS sorting and analysis, we used previously described staining protocols (Mossadegh-Keller et al., 2013 (link)), published stem and progenitor cell definitions (Bryder et al., 2006 (link)), FACSCanto, LSRII, and FACSAriaIII equipment, and DIVA software (BD), analyzing only populations with at least 200 events. The following antibodies were used for staining cells: anti-CD117 (clone 2B8; BD), anti–Sca-1 (clone D7; BioLegend), anti-CD34 (clone RAM34; BD), anti-CD16/32 (clone 2.4G2; BD), anti-CD11b (clone M1/70; BD), anti-CD19 (clone 1D3; BD), anti-CD3e (clone 145-2C11; BioLegend), anti-Ly6G (clone 1A8; BioLegend), anti-Ly6C (clone HK1.4; BioLegend), anti-CD115 (clone AFS98; eBioscience), anti-CD45.2 (clone 104; BD), anti-CD45.1 (clone A20; BD), anti-B220 (clone RA3-6B2; eBioscience), anti-Ter119 (clone TER-119; eBioscience), and anti-CD71 (clone R17217; eBioscience). LIVE/DEAD Fixable Violet Dead cell dye (Invitrogen) was used as a viability marker. Information describing gating strategies is shown in Figs. S1 and S2.

Kandalla P.K., Sarrazin S., Molawi K., Berruyer C., Redelberger D., Favel A., Bordi C., de Bentzmann S, & Sieweke M.H. (2016). M-CSF improves protection against bacterial and fungal infections after hematopoietic stem/progenitor cell transplantation. The Journal of Experimental Medicine, 213(11), 2269-2279.

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Multiplex Immunophenotyping of Murine Hematopoietic Cells

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The primary antibodies used were anti-CD3e (clone 145-2C11, catalog no. 100311, RRID:AB_312676), anti-CD11b (clone M1/70, catalog no. 101207, RRID:AB_312790, and catalog no. 101211, RRID: AB_312794), anti-CD27 (clone LG.3A10, catalog no. 124215, RRID:AB_10645330), anti-CD31 (clone 390, catalog no. 102409, RRID:AB_312904), anti-CD45 (clone 30-F11, catalog no. 103111, RRID:AB_312976), anti-CD45R (clone RA3-6B2, catalog no. 103211, RRID:AB_312996), anti-CD48 (clone HM48-1, catalog no. 103403, RRID:AB_313018), anti-CD51 (clone RMV-7, catalog no. 104105, RRID:AB_313074), anti-CD150 (clone TC15-12F12.2, catalog no. 115935, RRID:AB_2565960), anti-CD201 (clone RCR-16, catalog no. 141503, RRID:AB_10899579), anti-c-Kit (clone 2B8, catalog no. 105833, RRID:AB_2564054), anti-Gr-1 (clone RB6-8C5, catalog no. 108405, RRID:AB_313370, and catalog no. 108411, RRID:AB_313376), anti-Sca-1 (clone D7, catalog no. 108105, RRID:AB_313342), anti-TER-119 (clone TER-119, catalog no. 116211, RRID:AB_313712), and isotype-matched IgG controls conjugated with allophycocyanin (APC), FITC, PE, PE/Cyanine7 (Cy7), or PE/Dazzle, all of which were purchased from BioLegend. All other reagents used in this study were purchased from Sigma-Aldrich or FUJIFILM Wako Pure Chemical Corporation, unless otherwise specified.

Hiraga T., Ito S, & Mizoguchi T. (2021). Opposing Effects of Granulocyte Colony-Stimulating Factor on the Initiation and Progression of Breast Cancer Bone Metastases. Molecular cancer research : MCR, 19(12).

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Isolation and Identification of Spinal Cord Immune Cells

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Single-cell suspensions from spinal cord were obtained via mechanical dissociation on a cell strainer. Immune cells were separated over a two-phase Percoll-density gradient by centrifugation. Staining of CD4+ T cells, CD8+ T cells and CD45/CD11b+ cells (macrophages/microglia) was performed using the following antibodies in a 1:200 dilution: anti-CD3e (clone 145-2C11, BioLegend), anti-CD4 (clone GK 1.5, BD), anti-CD8 (clone 53-6.7, BD), anti-CD11b (clone M1/70, BioLegend), anti-CD45.2 (clone 104, BioLegend).
The addition of CaliBRITE APC beads (BD) allowed for cell quantification. Flow cytometry was performed using a CytoFLEX S (Beckman Coulter) operated by CytExpert software (Beckman Coulter, v2.4).

Berghoff S.A., Spieth L., Sun T., Hosang L., Depp C., Sasmita A.O., Vasileva M.H., Scholz P., Zhao Y., Krueger-Burg D., Wichert S., Brown E.R., Michail K., Nave K.A., Bonn S., Odoardi F., Rossner M., Ischebeck T., Edgar J.M, & Saher G. (2021). Neuronal cholesterol synthesis is essential for repair of chronically demyelinated lesions in mice. Cell reports, 37(4).

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Isolation and Quantification of Spinal Cord Immune Cells

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Single-cell suspensions from spinal cord were obtained via mechanical dissociation on a cell strainer. Immune cells were separated over a two-phase Percoll-density gradient by centrifugation. Staining of CD4+ T cells, CD8+ T cells and CD45/CD11b+ cells (macrophages/microglia) was performed using the following antibodies in a 1:200 dilution: anti-CD3e (clone 145-2C11, BioLegend), anti-CD4 (clone GK 1.5, BD), anti-CD8 (clone 53-6.7, BD), anti-CD11b (clone M1/70, BioLegend), anti-CD45.2 (clone 104, BioLegend). The addition of CaliBRITE APC beads (BD) allowed for cell quantification. Flow cytometry was performed using a CytoFLEX S (Beckman Coulter) operated by CytExpert software (Beckman Coulter, v2.4) .

Berghoff S.A., Spieth L., Sun T., Hosang L., Depp C., Sasmita A.O., Vasileva M.H., Scholz P., Zhao Y., Krueger‐Burg D., Wichert S.P., Brown E.R., Michael K., Nave K., Bonn S., Odoardi F., Rossner M.J., Ischebeck T., Edgar J.M., & Saher G. (2021). Neuronal cholesterol synthesis is essential for repair of chronically demyelinated lesions in mice.

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