Cd45ra pe

Manufactured by Miltenyi Biotec

CD45RA-PE is a laboratory reagent used for cell identification and analysis. It is a monoclonal antibody conjugated with the fluorescent dye R-Phycoerythrin (PE), which binds specifically to the CD45RA antigen expressed on the surface of certain human immune cells. This reagent can be used in flow cytometry applications to detect and quantify CD45RA-positive cells within a sample.

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CD45RA-PE-Vio770 (5 citations)

4 protocols using cd45ra pe

Multicolor Flow Cytometry Analysis of Immune Cells

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Cells were stained with CD3 FITC (Cat# 11-0039), CD3 eFluor 450 (Cat# 48-0038), CD4 PE-Cyanine7 (Cat# 25-0049), CD4 PerCP-Cyanine5.5 (Cat# 560650, BD Pharmingen), Foxp3 APC (Cat# 17-4777), Foxp3 PE (Cat# 12-4776), CD45RA PE (Cat# 130-092-248, Miltenyi), CD45RA APC (Cat# 130-092-249, Miltenyi), CD62L APC (Cat# 17-0629), CD25 PE (Cat# 130-091-024, Miltenyi), CD25 APC-H7 (Cat# 560225, BD Pharmingen), CD8 PE-Cyanine7 (Cat# 25-0088), Perforin PE (Cat# 12-9994), granzyme B PE (Cat# 12-8899), CCR8 APC (Cat# FAB1429A, R&D System), CD4 FITC (Cat# 11-0049), CD45 APC (Cat# 17-9459), CD14 PE (Cat# 12-0149), CD45RO PE (Cat# 12-0457), CD45RO eFluor 450 (Cat# 48-0457), CD127 APC (Cat# 17-1278). PITPNM3 antibody (Cat# NBP1-31070, Novus) was labeled with APC by Abcam APC Conjugation Kit (ab201807, Abcam) according to the manufacturer's instructions. For the intracellular stain, cells were pretreated with Intracellular Fixation and Permeabilization kit (Cat# 88-8824) according to the manufacturer's instructions. All the reagents were from eBioscience unless indicated otherwise. Cells were subsequently analyzed by multicolor flow cytometry (Gallios, Beckman Coulter, China).

Su S., Liao J., Liu J., Huang D., He C., Chen F., Yang L., Wu W., Chen J., Lin L., Zeng Y., Ouyang N., Cui X., Yao H., Su F., Huang J.D., Lieberman J., Liu Q, & Song E. (2017). Blocking the recruitment of naive CD4+ T cells reverses immunosuppression in breast cancer. Cell Research, 27(4), 461-482.

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Isolation of T Cell Subsets

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Following respirometry, cell suspensions were centrifuged and PBMC were resuspended in cell separation buffer (PBS, 0.5% bovine serum albumin, 2 mM EDTA; Miltenyi Biotec, Bergisch-Gladbach, Germany) for the separation of T cell subsets. Appropriate volumes of antibodies (CD4-APC [10 µl/10⁷ cells], CD8-FITC [10 µl/10⁷ cells], CD45RA-PE [5 µl/10⁷ cells], CD3-PE-Vio770 [5 µl/10⁷ cells], all purchased from Miltenyi Biotec) were added and cells were stained according to the manufacturer’s protocol. Naïve T helper cells (CD3⁺CD4⁺CD45RA⁺), memory T helper cells (CD3⁺CD4⁺CD45RA⁻), naïve cytotoxic T cells (CD3⁺CD8⁺CD45RA⁺), and memory cytotoxic T cells (CD3⁺CD8⁺CD45RA⁻) were then isolated by fluorescent-activated cell sorting on a BD FACSAria III cell sorter (BD Biosciences, Heidelberg, Germany). Propidium iodide staining was used to distinguish dead from living cells.

Boeck C., Salinas-Manrique J., Calzia E., Radermacher P., von Arnim C.A., Dietrich D.E., Kolassa I.T, & Karabatsiakis A. (2018). Targeting the association between telomere length and immuno-cellular bioenergetics in female patients with Major Depressive Disorder. Scientific Reports, 8, 9419.

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Immunophenotyping of PBMC and T-cells

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Staining of cell surface markers on PBMC and T-cells was performed with CD3-APC/Vio770, CD4-Vioblue, CD8-Viogreen, CD19-FITC, CD56-PE/Vio770, CD16-PE, CD62L-Vioblue, CD45RA-PE, CD45RO-APC, and CCR7-FITC (Miltenyi, San Diego, CA; BD, Franklin Lakes, NJ). All samples were acquired on a MACSQuant Cytometry (Miltenyi) and the data analyzed with Flow Jo (Treestar, Ashland, OR).

McLaughlin L.P., Lang H., Williams E., Wright K.E., Powell A., Cruz C.R., Colberg-Poley A.M., Barese C., Hanley P.J., Bollard C.M, & Keller M.D. (2016). Human Parainfluenza Virus-3 can be Targeted by Rapidly ex vivo Expanded T-Lymphocytes. Cytotherapy, 18(12), 1515-1524.

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Comprehensive CBMC and T Cell Immunophenotyping

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Staining of cell surface markers on CBMCs and T cells was performed with CD3-APC/Vio770, CD4-Vioblue, CD8-Viogreen, CD19-FITC, CD56-PE/Vio770, CD16-PE, CD62L-Vioblue, CD45RA-PE, CD45RO-APC, and CCR7-FITC (Miltenyi Biotec and BD Biosciences). 50,000 events per sample were acquired on a MACSQuant cytometer (Miltenyi), and the data were analyzed with Flow Jo (Tree Star).

Dave H., Luo M., Blaney J.W., Patel S., Barese C., Cruz C.R., Shpall E.J., Bollard C.M, & Hanley P.J. (2017). Toward a Rapid Production of Multivirus-Specific T Cells Targeting BKV, Adenovirus, CMV, and EBV from Umbilical Cord Blood. Molecular Therapy. Methods & Clinical Development, 5, 13-21.

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