ADSCs were characterized by flow cytometry at passage 4. Briefly, ADSCs were harvested and washed twice with phosphate-buffered saline (PBS). Then, the cells were incubated for 30 min in PBS containing anti-CD29-FITC (cat. # 555005; BD, San Diego, CA, USA), CD31-PE (cat. # 555027; BD, San Diego, CA, USA), CD49-FITC (cat. # 557457; BD, San Diego, CA, USA), CD90-PE (cat. # 551401; BD, San Diego, CA, USA), CD106-PE (cat. # 559229; BD, San Diego, CA, USA), CD34-FITC (cat. # sc-7324; Santa Cruz, Dallas, TX, USA), CD45-FITC (cat. # MCA43FT; AbD, Oxford, UK), CD73-FITC (cat. # bs-23233R; Bioss, Beijing, China), CD105-FITC (cat. # bs-10662R; Bioss, Beijing, China). The stained cells were then subjected to flow cytometry analysis.
Cd31 pe
Manufactured by BD
Sourced in United States, China
CD31-PE is a lab equipment product used for the detection and analysis of CD31-positive cells. It is a fluorescently labeled antibody that binds specifically to the CD31 cell surface antigen, also known as PECAM-1. The CD31-PE product can be used in various cell analysis and sorting applications, such as flow cytometry, to identify and quantify CD31-expressing cells within a sample.
Automatically generated - may contain errors
Lab products found in correlation
Cd45 fitc, by BD (16 mentions)
Cd73 pe, by BD (15 mentions)
Cd34 pe, by BD (11 mentions)
Cd34 fitc, by BD (10 mentions)
Cd44 pe, by BD (10 mentions)
Cd105 pe, by BD (9 mentions)
Cd90 pe, by BD (9 mentions)
Facscalibur flow cytometer, by BD (8 mentions)
Facscanto 2, by BD (7 mentions)
Cd90 fitc, by BD (6 mentions)
Cd45 pe, by BD (6 mentions)
Facscalibur, by BD (6 mentions)
Cd73 fitc, by BD (6 mentions)
Cd29 pe, by BD (6 mentions)
Cd45 apc, by BD (4 mentions)
Hla dr fitc, by BD (4 mentions)
Cd45 pe cy7, by BD (4 mentions)
Cd34 apc, by BD (4 mentions)
Cd146 pe, by BD (4 mentions)
Cd166 pe, by BD (4 mentions)
77 protocols using cd31 pe
1
Characterization of Adipose-Derived Stem Cells
2
Characterization of Cultured Adipose Stem Cells
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For flow cytometric analysis, cells were dissociated into single cell suspensions by incubation in trypsin/EDTA for 5 min. The dissociated cells were resuspended (1 × 105 cells) in 100uL of FACS buffer (PBS + 5% FBS + 0.1% Sodium azide), and then incubated for 30 minutes at 4 °C. After washing twice with FACS buffer, cells were analyzed with a fluorescence-activated cell sorter (FACS) (FACS Canto II, Becton Dickinson, San Jose, CA), and the acquired data were analyzed (Cell Quest software, Becton Dickinson, San Jose, CA). Cultured ASCs were characterized using antibodies against rat CD73 (BD Biosciences, 551123), CD90-FITC (BD Biosciences, 554897), CD31-PE (BD Biosciences, 555027), CD34 (Santa Cruz Biotechnology, sc-7324), and CD45-FITC (BD Biosciences, 554877). Anti-CD34 and CD73 antibodies were labeled with Alexa Fluor 488 (Thermo Fisher Scientific) Species-specific fluorophore (Alexa-Fluor 488) conjugated secondary antibodies (Abcam).
Lentiviral vector-transfected RLMVECs were analyzed by FACS with mouse anti-rat CD90-FITC and CD31-PE to determine whether they retained their vascular phenotype (Supplementary Fig.5 ).
Lentiviral vector-transfected RLMVECs were analyzed by FACS with mouse anti-rat CD90-FITC and CD31-PE to determine whether they retained their vascular phenotype (Supplementary Fig.
3
Flow Cytometry Analysis of CD34+ Cells
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Human nonexpanded and expanded PB-CD34+ cells (n = 5) were analyzed by flow cytometry (FACSCalibur flow cytometer, (Becton-Dickinson (BD), San Jose, CA). Dead cells were excluded by propidium iodide (PI) staining (Sigma, St Louis, MO). The CD34+ cells were incubated with a FcR blocking reagent (Miltenyi Biotec, Auburn, CA) and incubated with the monoclonal antibodies for 30 minutes at 4 °C. The stained cells were washed, resuspended, and then analyzed using Quad Statistics of CellQuest software (BD). The following monoclonal antihuman antibodies were used to characterized the CD34+ cell population: CD34-FITC (BD), CD31-PE (BD), CD133-PE (Miltenyi Biotec, Auburn, CA), CD68-PE (BD), CD83-PE (BD), VE-cadherin-PE (BD), VEGFR-2-PE (R&D Systems, Minneapolis, MN), Tie-2 (BD), CD117-PE (BD), CD45-PE (BD), IgG2a-FITC isotope controls (Miltenyi Biotec), and IgG1-PE isotope controls (Miltenyi Biotec).27 (link)The DNA content analysis was assessed by staining ethanol-fixed cells with PI and monitoring with the FACSCalibur flow cytometer. At least 20,000 cells were collected and analyzed with CellQuest software. Cell cycle distributions were calculated with ModFit LT cell-cycle analysis software (Verity Software House, Topsham, ME).
4
Comprehensive Antibody Panel for Cell Analysis
5
Comprehensive Immune Cell Phenotyping
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The following antibodies were used in this study: CD3 AF700, CD4 BV421, CD4 FITC, CD6 APC, CD10 BV605, CD19 APC-Cy7, CD21 PE-Cy7, CD25 PerCP-Cy5.5, CD27 BV421, CD28 PerCP-Cy5.5, CD38 PerCp-Cy5.5, CD45RA APC-Cy7, TCR αβ PerCP-Cy5.5, IL-2 PE, LAT PE, NTAL/LAB APC, and biotin anti–human TCR-Vd2 (BioLegend); TCR-Vd1 APC (Miltenyi Biotec); CD8 APC, CD8 Pacific blue, CD21 PE-Cy7, CD31 PE, CD56 APC, CD69 FITC, CD107a PE, CD127 Alexa Fluor 647, IFN-γ FITC, TCR γδ PE, IgG Alexa Fluor 700, IκBα PE, ERK1/2(pT202/pY204) AF647, and ZAP70(pY319)/SYK(pY352) APC (BD); IgD FITC and IgA PE (SouthernBiotech); CD3 PE-Cy7, CD4 PE-Cy7, CD8 PE, CD16 FITC, CD45RA FITC, CD45 Pacific blue, Vα24 PE, and Vβ11 FITC (Beckman Coulter); CCR7 PE (R&D Systems); Bruton tyrosine kinase/ITK(pY551/pY511) PE, IL-17 PE, IL-4 APC, and ICOS PE (eBioscience); IgM Alexa Fluor 647 (Jackson ImmunoResearch Laboratories, Inc.); and PLCγ1(pY783) and goat anti–rabbit AF647 (Cell Signaling Technology). For immunohistochemistry, CD21 (Dako), CD20 (Invitrogen), CD3 (Cell Marque), CD4 and CD8 (Spring Bioscience), and Bcl-6 (Leica Biosystems) were used. For immunoblotting LAT (sc-7948; Santa Cruz Biotechnology, Inc.), FLAG tag (AHP1074; AbD Serotec), actin (sc-1616; Santa Cruz Biotechnology, Inc.), PLCγ1 (pY783; no. 2821; Cell Signaling Technology), and ZAP70 (pY319; no. 2701; Cell Signaling Technology) were used.
6
Differentiation Marker Analysis of Embryoid Bodies
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EBs were dissociated with Collagenase IV in 37 °C and 5 % CO2 for 2 h. Single cell suspension from dissociated EBs was resuspended in 3 % FBS-PBS. The cell were passed through a 70 μm cell strainer and incubated at 4 °C for 1 h with the following fluorochrome-conjugated mouse anti-human antibodies: CD31-PE, CD34-FITC, CD45-APC, CD235a-PE, and CD71-APC (all BD Biosciences) or their corresponding isotype controls. Anti-OCT4 (BD Biosciences) and anti-ER-α (Santa Cruz) staining was identified using Alexa 488- and 647-conjugated goat anti-mouse IgG (Invitrogen). After washing with 3 % FBS-PBS, the cells were stained with 7-amino actinomycin to exclude dead cells. Flow analysis was performed on a FACSCanto II running BD FACSDiva™ (BD Biosciences) and acquired data were analyzed using FlowJo version 10 (Tree Star, Inc.).
7
Quantification of Endothelial and Microparticles
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Blood samples were obtained after 12 hours of fasting and the analyses were
performed at the central laboratory of our university. All athletes were
allowed to maintain their daily exercises program even on the day before
blood sample collection. The athletes had very similar exercise training
programs, corresponding to two long-distance running sessions every day, 15
km in the morning and 10 km in the afternoon, and intensive training
(100-1,000 meter shots, repeated many times)twice a week, on Tuesday and
Thursday mornings. All blood samples were collected on Thursdays, before
exercise.
Measurements of EPCs and MPs were performed as previously reported, using
fresh blood samples in EDTA containing tubes.12 (link)-15 (link)For determination of EPCs, a minimum of 500,000 events was acquired by
flow-cytometry (FACSCalibur, BD Biosciences, USA). Fluorescently labeled
mouse anti-human antibodies were used for EPCs (CD34 FITC, BD Biosciences,
USA; CD133 APC, Miltenyi Biotec, USA; KDR PE, R&D Systems, USA), PMPs
(CD42 FITC and CD31 PE, BD Biosciences, USA) and EMPs (CD51 FITC, BD
Biosciences). Disposable containers (BD Biosciences) were used to quantify
the number of microparticles per microliter of
platelet-poor plasma (PPP).
performed at the central laboratory of our university. All athletes were
allowed to maintain their daily exercises program even on the day before
blood sample collection. The athletes had very similar exercise training
programs, corresponding to two long-distance running sessions every day, 15
km in the morning and 10 km in the afternoon, and intensive training
(100-1,000 meter shots, repeated many times)twice a week, on Tuesday and
Thursday mornings. All blood samples were collected on Thursdays, before
exercise.
Measurements of EPCs and MPs were performed as previously reported, using
fresh blood samples in EDTA containing tubes.12 (link)-15 (link)For determination of EPCs, a minimum of 500,000 events was acquired by
flow-cytometry (FACSCalibur, BD Biosciences, USA). Fluorescently labeled
mouse anti-human antibodies were used for EPCs (CD34 FITC, BD Biosciences,
USA; CD133 APC, Miltenyi Biotec, USA; KDR PE, R&D Systems, USA), PMPs
(CD42 FITC and CD31 PE, BD Biosciences, USA) and EMPs (CD51 FITC, BD
Biosciences). Disposable containers (BD Biosciences) were used to quantify
the number of microparticles per microliter of
platelet-poor plasma (PPP).
8
Immunophenotyping of Cultured Cells
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After cell detachment using a 0.125% trypsin solution, cells were washed with PBS and resuspended in PBS containing 2% FBS. Cell concentration and viability were monitored using Trypan blue in a Neubauer haemocytometer. The following monoclonal antibodies were used as indicated by the manufacturer (BD Pharmingen): CD90-PE (BD, #555596), CD73-FITC (BD, #561254), CD105-FITC (BD,#561443), CD45-FITC (BD,#347463), CD14-PE (BD,#555398), CD34-PEcy5 (BD,#561819), CD31-PE (BD,#555446), IgG-FITC (BD,#555786), HLA-DR-FITC (BD,#555558), CD166-PE (BD,#560903), CD44-PE (BD,#555479), CD54-PEcy5 (BD,#555512), CD146-PE (BD,# 559263). At least 20,000 events were acquired on a BD FACSCalibur flow cytometer and data was analyzed using CellQuest software.
9
Phenotypic Characterization of MSCs
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After the fourth passage, MSCs were detached
by trypsinization. The cells were washed with
PBS, and incubated with labeled phycoerythrin
(PE)-conjugated monoclonal antibodies (Invitrogen,
USA) at the dilutions recommended
by the manufacturer at 4˚C for 25 minutes in
the dark. The antibodies used were cluster of
differentiation (CD) markers; CD90, CD34,
CD105, and CD31-PE (BD Biosciences, USA).
PE-labeled isotype-matched immunoglobulin
was the negative control. The labeled cells
were analyzed on a FACS Caliber flow cytometer
(Becton-Dickinson, FACScan, San Jose,
CA, USA).
by trypsinization. The cells were washed with
PBS, and incubated with labeled phycoerythrin
(PE)-conjugated monoclonal antibodies (Invitrogen,
USA) at the dilutions recommended
by the manufacturer at 4˚C for 25 minutes in
the dark. The antibodies used were cluster of
differentiation (CD) markers; CD90, CD34,
CD105, and CD31-PE (BD Biosciences, USA).
PE-labeled isotype-matched immunoglobulin
was the negative control. The labeled cells
were analyzed on a FACS Caliber flow cytometer
(Becton-Dickinson, FACScan, San Jose,
CA, USA).
10
Endothelial Cell Surface Marker Analysis
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The cells were harvested by 0.05% trypsin treatment for 3 min, then the cells were washed once with FACS buffer (0.5% BSA/PBS) and fixed with 1% PFA at 37°C for 10 min followed by washing 3 times. The CD31-PE (1:100, BD), CD144-FITC (1:100, Bioss, China) antibodies were diluted in FACS buffer and incubated with the cells at 4°C for 30 min. After 3 times washing with FACS buffer, the cells were analyzed on the Cytoflex LX flow cytometer (Beckman, USA).
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