Anti cd16fitc

Manufactured by Beckman Coulter

Anti-CD16 FITC is a fluorescently labeled antibody that binds to the CD16 antigen expressed on the surface of certain immune cells, such as natural killer cells and a subset of monocytes. It is commonly used in flow cytometry applications to identify and quantify these cell populations.

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3 protocols using anti cd16fitc

Multiparameter Flow Cytometry of PBMCs

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After separation from the whole blood in EDTA, 100 μl of viable PBMC (peripheral blood mononuclear cell) was stained with different combinations of monoclonal antibodies. To characterize the phenotype of T and B cell subsets, extracellular labeling was performed with anti-CD8_FITC, anti-CD161_PE, anti-CD3_ECD, anti-CD4_PC5.5, anti-CD16_FITC, anti-CD56_PE, anti-IgD_FITC, anti-CD27_PC5.5, and anti-CD19_ECD (Beckman Coulter, Miami, FL). Living cells were gated within the side/forward scatter (SSC/FSC) lymphocyte gate. For intracellular staining, cells were permeabilized with Cytofix/Citoperm (BD Biosciences). Finally, the cells were stained with anti-IL-17A_FITC (MiltenyiBiotec), washed, and analyzed. All measurements were made with a CyAN ADP flow cytometer (Beckman Coulter, Miami, FL, USA) with the same instrument setting. At least 105 lymphocytes were acquired and analyzed using FlowJo (Tree Star) software. Leukocyte count and differential were determined with a routine hematology analyzer. The absolute counts of total lymphocytes were calculated by multiplying the relative size of the T and B cells and the absolute lymphocyte count.

Balistreri C.R., Buffa S., Allegra A., Pisano C., Ruvolo G., Colonna-Romano G., Lio D., Mazzesi G., Schiavon S., Greco E., Palmerio S., Sciarretta S., Cavarretta E., Marullo A.G, & Frati G. (2018). A Typical Immune T/B Subset Profile Characterizes Bicuspid Aortic Valve: In an Old Status?. Oxidative Medicine and Cellular Longevity, 2018, 5879281.

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Multiparameter Flow Cytometry of Immune Cell Markers

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The following antibodies were used: anti CD14-BV510, anti CD3–BV421 and anti CD56–PECy7 from BD Biosciences; anti CD300lf-PE from BD Biosciences or anti CD55-APC from Biolegend; anti CD16-APC from BD Biosciences or anti CD16-FITC from Beckman Coulter (Miami, FL) and PE Mouse IgG1, κ Isotype Control from BD Biosciences or APC Mouse IgG1, κ Isotype Control from R&D System. Red blood cell lysis was performed using Versalyse lysing solution (Beckman Coulter). CD300LF and CD55 expression were measured using Navios flow cytometer (Beckman-Coulter). Results were analyzed with Kaluza software (Beckman-Coulter) expressed as Medians of Fluorescence Intensity (MFI).

Tabone O., Mommert M., Jourdan C., Cerrato E., Legrand M., Lepape A., Allaouchiche B., Rimmelé T., Pachot A., Monneret G., Venet F., Mallet F, & Textoris J. (2019). Endogenous Retroviruses Transcriptional Modulation After Severe Infection, Trauma and Burn. Frontiers in Immunology, 9, 3091.

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Flow Cytometry for NK Cell Analysis

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Multi-parameter flow cytometry was performed using a lysed-whole-blood technique without isolation of cells on a density gradient, using a commercially available red cell lysing solution, Optilyse B (Beckman Coulter). The cell labelling and membrane fixation/permeabilization procedures were done by standard methods; the cell staining was performed as described in (Babusikova et al. 2008) . We used monoclonal antibodies targeting membrane antigens: anti-CD45-ECD (Beckman Coulter), anti-CD16-FITC (Beckman Coulter), anti-CD11b-PE (Miltenyi Biotec), anti-CD3-FITC (Miltenyi Biotec), anti CD4-PE (Miltenyi Biotec), anti-CD8-APC (Miltenyi Biotec), anti-CD25-Pacific Blue (ExBio Praha), anti-CD19-FITC (Miltenyi Biotec) and anti-CD56-PE (Beckman Coulter), and FACS analysis was performed by FACS Canto II flow cytometer (BD Biosciences, San Jose, CA). The data were analysed using FCS Express 4.0 (De Novo Software). NK cells were identified as CD3-CD56 + cells. To determine the absolute number of NK cells, equal volumes (100 μl) of Flow-Count Fluorospheres (Beckman Coulter, CA, USA) and cells were mixed, and the NK cell concentration was calculated according to the formula: NK cells/μl = (count NK cells × concentration beads)/ (count beads).

Gronesova P., Cholujova D., Kozic K., Korbuly M., Vlcek M., Penesova A., Imrich R., Sedlak J, & Hunakova L. (2018). Effects of short-term Pilates exercise on selected blood parameters. General physiology and biophysics, 37(4).

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