After 10 passages, the cells were characterized by multiple commonly utilized markers for conventional MSCs. Phenotypic antibodies used are as follows: CD45-FITC, CD90-FITC, CD34-PE, CD11c-FITC, CD44-PE, I-Ab-FITC, CD80-FITC, CD11b-PE-Cy7, CD73-PE (all from BD Biosciences) and H-2kb-APC, CD105-PE, Sca-1-FITC, CD9-PE, and CD86-PE (all from eBioscience). All isotype controls were purchased from BioLegend. Flow Cytometry was conducted on a BD FacsCalibur. Standard protocols were conducted to verify the ability of MSCs to differentiate into adipocytes and osteoblasts using Mesencult mouse adipogenic stimulatory supplements and mouse osteogenic stimulatory supplements (StemCell Technologies products), respectively, in IMDM-based media with 10% FBS supplementation. Supplement-to-basal media was at a 1:4 ratio. For both differentiation tests, 2.5×104 MSC were plated per well in 2mL Mesencult media in 6-well plates in 37°C/ 10% CO2 incubation. On day 3, cells were given appropriate differentiation media, with media changes every 3– 4 days. After 2 weeks of culture, cells were stained for adipocytic differentiation with Oil Red O and osteogenic differentiation with Alizarin red. Human MSCs were phenotyped by Lonza as CD105+CD166+CD29+CD44+ > 90% and CD14+CD34+CD45+ <10%.
I ab fitc
Manufactured by BD
Sourced in France, United Kingdom
I-Ab-FITC is a fluorescently-labeled secondary antibody used for detection in immunoassays. The FITC (fluorescein isothiocyanate) label allows for visualization of the target analyte.
Automatically generated - may contain errors
Lab products found in correlation
Cd11c fitc, by BD (2 mentions)
Facscalibur, by BD (1 mentions)
Sca 1 fitc, by Thermo Fisher Scientific (1 mentions)
Cd44 pe, by BD (1 mentions)
Cd86 pe, by Thermo Fisher Scientific (1 mentions)
Cd105 pe, by Thermo Fisher Scientific (1 mentions)
Cd45 fitc, by BD (1 mentions)
Cd34 pe, by BD (1 mentions)
Cd80 fitc, by BD (1 mentions)
Cd73 pe, by BD (1 mentions)
Cd90 fitc, by BD (1 mentions)
Cd11b pe cy7, by BD (1 mentions)
H 2kb apc, by Thermo Fisher Scientific (1 mentions)
Cd11b apc cy7, by BD (1 mentions)
Cd4 pe cy7, by BD (1 mentions)
Cd19 apc, by BD (1 mentions)
Cd11c pe cy7, by BD (1 mentions)
Lipopolysaccharides (lps), by Merck Group (1 mentions)
Interleukin 6 (il 6), by Merck Group (1 mentions)
Cd45.1 apc, by BD (1 mentions)
3 protocols using i ab fitc
1
Characterization of Mesenchymal Stem Cells
2
Murine Immune Cell Phenotyping
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Murine GM-CSF was from Peprotech (Neuilly-sur-Seine, France). IL-6 and LPS were from Sigma-Aldrich (Saint-Quentin Fallavier, France). CFDA-SE (CFSE) was from Molecular Probes (Montluçon, France). OVA (SIINFEKL) and Smcy (KCSRNRQYL) peptides were from PolyPeptide (Strasbourg, France). Anti mouse CD11b biotin (M1/70) (used with streptavidin APC or streptavidin APC-Cy7), CD11b APC-Cy7 (M1/70), CD11c PE-Cy7 (HL3), I-Ab FITC (AF6-120.1), Gr1 PE (Ly6C/G, RB6-8C5), CD45.1 APC (A20), CD45.2 APC-Cy7 (104), CD45.2 PerCP-Cy5.5 (104), CD19 APC (1D3), NK1.1 PE (PK136), CD3ε PerCP-Cy5.5 (145-2C11), CD3ε Pacific Blue (500A2), CD3ε FITC (145-2C11), CD4 PE-Cy7 (RM4-5), CD8α Pacific blue (53-6.7), CD8α APC-Cy7 (53-6.7), CD8α PerCP-Cy5.5 (53-6.7), FoxP3 Alexa Fluor647 (MF23), CD25 PE (704), CD69 FITC (H1.2F3), and CD86 FITC (B7.2, GL1) were from BD PharMingen (Le Pont de Claix, France). Male antigen UTY-specific CD8+ T cells were detected using a PE labelled Pro5 MHC Pentamer (H-2Db, WMHHNMDLI) (ProImmune Limited, Oxford, UK).
3
Cardiac Inflammatory Cell Profiling after MI
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From each treatment group, mice not subjected to functional and histological analysis were used for flow cytometric analysis of cardiac inflammatory cell invasion. Mice were sacrificed on days 1, 3, and 7 after MI. Each time point, 3 hearts were harvested per group. Sham animals were used as controls to determine base line characteristics. Total cardiac cell numbers were determined with a Sysmex cell counter (Sysmex America, Inc. Mundelein, Illinois, US). Single-cell suspensions were stained for flow cytometry with primary antibodies before analysis using a FACSCanto II (BD Biosciences, San Diego, CA, USA). The following antibodies were used: anti–CD90-APC, 53–2.1,–B220-APC, RA3-6B2,–CD49b-APC, DX5,–NK1.1-APC, PK136,–Ly-6G-APC, 1A8, CD11b-eFluor 450, M1/70,–CD11c-FITC, HL3,–I-Ab -FITC, AF6-120.1,–Ly-6C-PE, AL-21,–CD11c-PE, HL3 (All above antibodies are from BD Biosciences),–F4/80-FITC, C1:A3-1 (ABD Serotec, Kidlington, UK). Monocytes were identified as CD11b high (CD90/B220/CD49b/NK1.1/Ly-6G) low (F4/80/I-Ab /CD11c) low Ly-6C high/low as previously described [26 (link),28 (link)]. Macrophages were identified as CD11b high, F4/80 high. Dendritic cells were identified as CD11b, I-AbandCD11c high. Neutrophils were identified as CD11b, Ly-6G high. The analysis of the acquired data was done with FlowJo software version 7.6.1 (Tree Star Inc. Ashland, OR, USA).
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