Cd45.2 fitc 104

Manufactured by BioLegend

CD45.2 FITC (104) is a fluorochrome-conjugated antibody that recognizes the CD45.2 isoform of the CD45 antigen. CD45 is a family of transmembrane tyrosine phosphatases expressed on the surface of most hematopoietic cells. The CD45.2 isoform is specifically found on cells of mouse origin.

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4 protocols using cd45.2 fitc 104

Comprehensive Immune Cell Profiling

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The following antibodies were used (all from BioLegend): CD3 (clone 17A2), Ter119 (TER-119), Gr-1 (RB6–8C5), B220 (RA3–6B2) conjugated to Pacific Blue, CD117 APC-Cy7 (2B8), Sca-1 PE-Cy7 (D7), Flk2 PE (A2F10), CD48 FITC (HM48–1), CD150 APC (TC15–12F12.2), B220 PE-Cy7 (RA3–6B2), Mac-1 PE-Cy5 (M1/70), CD3 PE (17A2), Ter119 APC-Cy7 (TER-119), CD45.2 FITC (104), CD45.1 APC-Cy7 (A20), CD16/32 PerCP-Cy5.5 (93), CD5 APC-Cy7 (53–7.3). CD19 PE (eBio1D3) and CD34 FITC (RAM34) were from eBioscience. Cells were labeled with SYTOX Blue viability stain (Thermo). Data were acquired on LSRII or LSR Fortessa instruments (BD Biosciences). APC-annexin V was from BioLegend.

Wang D., Tanaka-Yano M., Meader E., Kinney M.A., Morris V., Lummertz da Rocha E., Liu N., Liu T., Zhu Q., Orkin S.H., North T.E., Daley G.Q, & Rowe R.G. (2022). Developmental maturation of the hematopoietic system controlled by a Lin28b-let-7-Cbx2 axis. Cell reports, 39(1), 110587.

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Comprehensive Immune Cell Profiling

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Multicolor Flow Cytometry for Immune Cell Profiling

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For flow cytometric analysis, the directly labeled mAbs used were CD11b-PerCP (M1/70), CD4-APC-Cy7 (L3T4) and Ly-6G-PE-Cy7 (1A8) from Biolegend, CD45.2-FITC (104), CD11c-APC (HL3), NK1.1-PE (PK136), Ly-6C-APC-Cy7 (AL-21) and CD3-PE (145-2C11) from BD, MHC II-PE (M5/114.15.2) from eBioscience. Cells for staining were washed in PBS and incubated with Fixable Viability Dye eFluor 780 (eBioscience). Cells were resuspended in ice-cold PBA and transferred to a V-bottom 96-wells plate. After incubation for 30 min with PBA containing anti-CD16/CD32 Fc-block (BD), cells were stained using specific monoclonal antibodies or the appropriate isotype controls for a period of 30 min. Cells were washed in PBA and measured on a Cyan apparatus (BD) and analyzed using Summit software.

Kroesen M., Brok I.C., Reijnen D., van Hout-Kuijer M.A., Zeelenberg I.S., Den Brok M.H., Hoogerbrugge P.M, & Adema G.J. (2015). Intra-adrenal murine TH-MYCN neuroblastoma tumors grow more aggressive and exhibit a distinct tumor microenvironment relative to their subcutaneous equivalents. Cancer Immunology, Immunotherapy, 64(5), 563-572.

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Thymus Tissue Immunofluorescence Staining

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Thymus grafts were embedded in OCT Compound and snap-frozen in liquid nitrogen. Tissue sections of 8 mm were dehydrated in acetone and samples were preserved in a dry environment at À80 C until staining. Sections were incubated for 30 min with DAPI and mIgG in PBS/10%FBS at room temperature. Following two washes in PBS the samples were stained with CD25 PE (PC61), CD45.2 FITC (104) and Cytokeratin 8 A647 (1E8) overnight at 4 C. Antibodies were purchased from Biolegend. Stained sections were washed three times in PBS and mounted with Fluoromount-G (Thermo Fisher Scientific). Images were acquired on a commercial Nikon High Content Screening microscope, equipped with an Andor Zyla 4.2 sCMOS camera, using a 20x 0.75 NA objective.

Ramos C.V., Ballesteros-Arias L., Silva J.G., Paiva R.A., Nogueira M.F., Carneiro J., Gjini E, & Martins V.C. (2020). Cell Competition, the Kinetics of Thymopoiesis, and Thymus Cellularity Are Regulated by Double-Negative 2 to 3 Early Thymocytes. Cell reports, 32(3).

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