Clone fn50

Manufactured by BD

Sourced in United States

The Clone FN50 is a laboratory instrument designed for the detection and quantification of fluorescent signals. It features a compact and efficient optical system that allows for the measurement of fluorescence intensity in a variety of sample types, including microplates, cuvettes, and microscope slides. The Clone FN50 is suitable for a range of applications in fields such as molecular biology, biochemistry, and cell biology.

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Lab products found in correlation

Amcyan, by Thermo Fisher Scientific (2 mentions) Clone l200, by BD (2 mentions) Anti cd4 v50, by BD (2 mentions) Clone dk25, by Agilent Technologies (2 mentions) Clone l48, by BD (2 mentions) Clone 4b4 1, by Miltenyi Biotec (2 mentions) Clone l106, by BD (2 mentions) Facslyric, by BD (1 mentions) Clone mopc 21, by BioXCell (1 mentions) Cd16 apc cy7, by BD (1 mentions) Interleukin 2 (il 2), by Thermo Fisher Scientific (1 mentions) Clone cd28, by BD (1 mentions) Efluor 670, by BD (1 mentions) Clone rpa t4, by BD (1 mentions) Clone okt 3, by BioXCell (1 mentions) Facs lsr 2, by BD (1 mentions) Fortessa, by BD (1 mentions) Efluor 780, by Thermo Fisher Scientific (1 mentions) Cd20 apc clone l27, by BD (1 mentions) Cd8a bv785, by BioLegend (1 mentions)

Close spelling variants of this product

CD69 (clone FN50, (7 citations) CD69-PE (clone FN50), (6 citations) CD69-PerCP-Cy5–5 (clone FN50, (3 citations) Anti-CD69 (clone FN50, (2 citations) Anti CD69 PE (clone FN50), (2 citations) CD69-PE.Cy7 (clone FN50; (2 citations)

4 protocols using clone fn50

Multiparametric Flow Cytometry for SARS-CoV-2-Specific T-Cell Detection

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Detection of AIM was performed as described previously (8 (link)). Briefly, cells were stained with the following antibodies in their respective dilutions: anti–CD3-PerCP (1:25, clone SK7, BD), anti–CD4-V50 (1:50, clone L200; BD), anti–CD8–fluorescein isothiocyanate (1:25, clone DK25; Dako), anti–CD45RA-phycoerythrin (PE)–Cy7 (1:50, clone L48; BD), anti–CCR7-BV711, anti–CD69-allophycocyanin (APC)-H7 (1:50, clone FN50; BD), anti–CD137-PE (1:50, clone 4B4-1; Miltenyi), and anti–OX40-BV605 (1:25, clone L106; BD). LIVE/DEAD Fixable Aqua Dead Cell staining was included (1:100, AmCyan; Invitrogen) and acquired on a FACSLyric (BD Biosciences). After setting a time gate, LIVE CD3+ T-cells were gated, singlets were selected, and T-cells were subtyped into CD3+CD4+ and CD3+CD8+ cells. Within the CD4+ and CD8+ T-cells, T_naive were defined as CD45RA+CCR7+, T_CM as CD45RA-CCR7+, T_EM as CD45RA-CCR7-, and T_EMRA as CD45RA+CCR7-. S-specific T-cells were detected by co-expression of AIM on CD4+ (OX40 and CD137) or CD8+ (CD69 and CD137) T-cells in the combination of memory subsets (Fig. 3A). The DMSO-stimulated sample was used to set the cutoff gate for activation markers. On average, 500,000 cells were acquired per sample.

GeurtsvanKessel C.H., Geers D., Schmitz K.S., Mykytyn A.Z., Lamers M.M., Bogers S., Scherbeijn S., Gommers L., Sablerolles R.S., Nieuwkoop N.N., Rijsbergen L.C., van Dijk L.L., de Wilde J., Alblas K., Breugem T.I., Rijnders B.J., de Jager H., Weiskopf D., van der Kuy P.H., Sette A., Koopmans M.P., Grifoni A., Haagmans B.L, & de Vries R.D. (2022). Divergent SARS CoV-2 Omicron-reactive T- and B cell responses in COVID-19 vaccine recipients. Science Immunology.

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Activated T Cell Phenotyping by Flow Cytometry

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Flow cytometry was performed on a FACS LSR II or Fortessa (BD Biosciences). For lymphocyte activation studies, soluble anti-CD3 (500 ng/ml; clone OKT3, Bio X Cell) ± anti-CD28 (1 μg/ml; clone CD28.2, BD Biosciences) or IL-2 (PeproTech) was added to media for 2 days. Proliferation was measured by CFSE (eFluor 670, BD Biosciences) after 3 days. EVs were added every 24 hours (5 μg/ml). PD1 blockade was achieved by adding anti-PD1 (10 μg/ml; EH12) or immunoglobulin G control (clone MOPC-21, Bio X Cell). Cells were gated by FSC (forward scatter)/SCC (side scatter) while excluding duplets by FSC-A/FSC-H. Subsequently, CD3⁺CD56⁻ T cells were gated with further classification of CD4⁺ and CD8⁺ T cells. Finally, CD4⁺ and CD8⁺ T cells were measured for their CD69, CD25, PD1, and TIM3 expression (seen as a downloadable figure). Cells were stained with the following antibodies: CD3/PE-Cy5.5 (1:100; clone SK7, eBioscience), CD4/FITC (1:100; clone RPA-T4, BD Biosciences), CD8/AmCyan (1:10; clone SK1, BD Biosciences), CD16/APC-Cy7 (1:100, BD Biosciences), CD25/PE-Cy7 (1:10; clone BC96, eBioscience), CD56/V450 (1:30; clone B159, BD Biosciences), CD69/APC (1:10; clone FN50, BD Biosciences), TIM3/APC-Cy7 (1:100, clone F38-2E2, BD Biosciences), and PD1/PE (1:100; clone J105, eBioscience).

Ricklefs F.L., Alayo Q., Krenzlin H., Mahmoud A.B., Speranza M.C., Nakashima H., Hayes J.L., Lee K., Balaj L., Passaro C., Rooj A.K., Krasemann S., Carter B.S., Chen C.C., Steed T., Treiber J., Rodig S., Yang K., Nakano I., Lee H., Weissleder R., Breakefield X.O., Godlewski J., Westphal M., Lamszus K., Freeman G.J., Bronisz A., Lawler S.E, & Chiocca E.A. (2018). Immune evasion mediated by PD-L1 on glioblastoma-derived extracellular vesicles. Science Advances, 4(3), eaar2766.

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Isolation and Characterization of Brain-Derived Immune Cells

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Peripheral blood mononuclear cells and subcortical white matter‐derived single cell fractions were isolated after rapid post‐mortem autopsies of NBB brain donors as described previously 56, 57. Donor and sample characteristics are listed in Table S7. Cells were stained with fixable viability dye eFluor 780 (Life Technologies) and the following antibodies: CD3 PE‐Cy5.5, clone SK7, (Invitrogen), CD20 APC, clone L27, CD25 FITC, clone 2A3, CD69 BV395, clone FN50, CD127 PE, clone HIL‐7R‐M21, (BD Biosciences), CD4 BV510, clone RPA‐T4, CD8a BV785, clone RPA‐T8, and FAS PE‐Cy7, clone DX2, (Biolegend), and analyzed on a Fortessa LSR^TM cell analyzer (BD Biosciences, San Jose, CA, USA). Data were analyzed with FlowJo software 10.5 (Tree Star, Ashland, OR, USA).

Fransen N.L., Crusius J.B., Smolders J., Mizee M.R., van Eden C.G., Luchetti S., Remmerswaal E.B., Hamann J., Mason M.R, & Huitinga I. (2019). Post‐mortem multiple sclerosis lesion pathology is influenced by single nucleotide polymorphisms. Brain Pathology, 30(1), 106-119.

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Identifying SARS-CoV-2-specific T cells

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After ex vivo stimulation of PBMCs, cells were stained at 4°C for 15 min for phenotypical lymphocyte markers and AIM expression. Surface staining was performed with the following antibodies in their respective dilutions: anti–CD3-PerCP (1:25, clone SK7, BD), anti–CD4-V50 (1:50, clone L200; BD), anti–CD8–fluorescein isothiocyanate (1:25, clone DK25; Dako), anti–CD45RA-phycoerythrin (PE)–Cy7 (1:50, clone L48; BD), anti–CCR7-BV711, anti–CD69-allophycocyanin (APC)-H7 (1:50, clone FN50; BD), anti–CD137-PE (1:50, clone 4B4-1; Miltenyi), and anti–OX40-BV605 (1:25, clone L106; BD). LIVE/DEAD Fixable Aqua Dead Cell staining was included (1:100, AmCyan; Invitrogen). Cells were first gated for LIVE CD3⁺ T cells and then subdivided into CD3⁺CD4⁺ T helper cells and CD3⁺CD8⁺ T-cytotoxic cells (Fig. 4A). SARS-CoV-2–specific T cells were identified by gating the CD69⁺CD137⁺ cells (within CD4⁺ or CD8⁺ subsets). For N = 11 of 20 donors assessed, SARS-CoV-2 specificity in the CD4⁺ subset was confirmed by gating CD137⁺OX40⁺ cells. The DMSO-stimulated sample was used to set the cutoff gate for activation markers. On average, 500,000 cells were acquired per sample. Low-frequency samples (<10,000 cells in CD4 gate and <5000 cells in CD8 gate) were excluded from analysis.

Geers D., Shamier M.C., Bogers S., den Hartog G., Gommers L., Nieuwkoop N.N., Schmitz K.S., Rijsbergen L.C., van Osch J.A., Dijkhuizen E., Smits G., Comvalius A., van Mourik D., Caniels T.G., van Gils M.J., Sanders R.W., Oude Munnink B.B., Molenkamp R., de Jager H.J., Haagmans B.L., de Swart R.L., Koopmans M.P., van Binnendijk R.S., de Vries R.D, & GeurtsvanKessel C.H. (2021). SARS-CoV-2 variants of concern partially escape humoral but not T cell responses in COVID-19 convalescent donors and vaccine recipients. Science Immunology, 6(59), eabj1750.

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