Samples were incubated in 250 μl of 2% FBS/PBS with fluorochrome-conjugated primary antibodies for 30 min, with shaking every 10 min. Primary antibodies were washed with 2% FBS/PBS, and cells were resuspended in 2.5 mg ml−1 DAPI (Invitrogen, D1306) before analysis. The following primary antibodies were used: APC-conjugated anti-CD45 (1:100, clone 30-F11, eBiosciences), APC-conjugated anti-CD31 (1:100, clone 390, eBiosciences), APC-conjugated anti-CD140a (1:100, clone APA5, eBiosciences), PE-Cy7-conjugated anti-CD24 (1:100, clone M1/69, BD Biosciences), FITC-conjugated anti-CD29 (1:100, clone Ha2/5, BD Biosciences), APC-Cy7-conjugated anti-EpCAM (1:100, clone G8.8, BioLegend). Data analysis and cell sorting were performed on a FACSAria sorter using the FACS DiVa software (BD Biosciences). Dead cells were excluded with DAPI; CD45+, CD31+ and CD140a+ cells were excluded (Lin+) before analysis of the tdTomato+ cells.
Fitc conjugated anti cd29
Manufactured by BD
Sourced in United States
FITC-conjugated anti-CD29 is a fluorescently labeled antibody that specifically binds to the CD29 cell surface marker. CD29 is also known as the integrin beta-1 subunit and plays a role in cell-cell and cell-extracellular matrix interactions. This antibody can be used to identify and isolate cells expressing CD29 in flow cytometry and other applications.
Automatically generated - may contain errors
Lab products found in correlation
Apc conjugated anti cd45, by Thermo Fisher Scientific (2 mentions)
Apc conjugated anti cd31, by Thermo Fisher Scientific (2 mentions)
Apc conjugated anti cd140a, by Thermo Fisher Scientific (2 mentions)
Pecy7 conjugated anti cd24, by BD (2 mentions)
Facsdiva software, by BD (2 mentions)
Pe conjugated anti cd45, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti cd31, by BD (1 mentions)
Apc conjugated anti cd29, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti cd140a, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Lsrfortessa, by BD (1 mentions)
Cytoflex s flow cytometer, by Beckman Coulter (1 mentions)
Staining buffer, by BD (1 mentions)
Pe conjugated anti cd29, by Thermo Fisher Scientific (1 mentions)
Fcs express 6, by De Novo Software (1 mentions)
Pe conjugated anti cd61, by BD (1 mentions)
Anti tet2, by Cell Signaling Technology (1 mentions)
Facsaria, by BD (1 mentions)
Pe conjugated anti cd90, by BD (1 mentions)
Apc conjugated anti cd31, by BD (1 mentions)
5 protocols using fitc conjugated anti cd29
1
Fluorescence-Activated Cell Sorting of Lineage-Negative Cells
2
Multicolor Flow Cytometry Analysis of Murine Cell Lineages
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Two million to 5 million cells per condition were incubated in 250 µL of 2% FBS/PBS with fluorochrome-conjugated primary antibodies for 30 min with shaking every 10 min. Primary antibodies were washed with 2% FBS/PBS, and cells were resuspended in 2.5 mg/mL DAPI (Invitrogen) before analysis. The following primary antibodies were used for the analysis of K14rtTA/TetO-Cre/Rosa-YFP mice: PECy7-conjugated anti-CD24 (1:100; BD Biosciences, clone M1/69), APC-conjugated anti-CD29 (1:100; eBiosciences, clone eBioHMb1-1), PE-conjugated anti-CD45 (1:100; eBiosciences, clone 30-F11), PE-conjugated anti-CD31 (1:100; BD Biosciences, clone MEC 13.33), and PE-conjugated anti-CD140a (1:100; eBiosciences, clone APA5). Data analysis was performed on a BD LSR Fortessa using FACS DiVa software (BD Biosciences). Dead cells were excluded with DAPI; CD45+, CD31+, and CD140a+ cells were excluded (Lin+) before analysis of the YFP+ cells. Primary antibodies used for the analysis of K8rtTA/TetO-Cre/Rosa-tdTomato mice were PECy7-conjugated anti-CD24 (1:100; BD Biosciences, clone M1/69), FITC-conjugated anti-CD29 (1:100; BD Biosciences, clone Ha2/5), APC-conjugated anti-CD45 (1:100; eBiosciences, clone 30-F11), APC-conjugated anti-CD31 (1:100; eBiosciences, clone 390), and APC-conjugated anti-CD140a (1:100; eBiosciences, clone APA5). A minimum of five 105 total events per mouse were analyzed.
3
Brain Cell Isolation and Flow Cytometry
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Mice were sacrificed and the brains were rapidly dissected out. After mincing into 1–2 mm pieces, the cerebral hemispheres were incubated for 30 min at 37 ˚C in the presence of 10 unit/ml papain and 0.2 mg/ml DNase. Following filtration through a 70-μm pore size nylon mesh and removal of the papain solution, 30% Percoll was added and the cell suspension was centrifuged for 30 min at 500×g. The cell pellet was used for the Flow cytometry analyses. Cells were stained with the following antibodies: APC conjugated anti-ACSA-2 (Miltenyi Biotec, Cat#130-116-245, clone REA969, 1:100) and FITC conjugated anti-CD29 (BD Biosciences, Cat#102205, clone HMβ1-1, 1:100). Cells were analyzed with Cytoflex S Flow cytometer (Beckman Coulter) and data were analyzed with FlowJo software version 10.8.1 (BD Biosciences).
4
Multiparametric Flow Cytometry Analysis
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Cells at a concentration of 1 × 106 per 100 µl of staining buffer (BD Biosciences) were incubated on ice for 30 min with the following antibodies: FITC-conjugated anti-CD29 (#561796, BD Biosciences, 1:100), PE-Cy7-conjugated CD29 (#25029182, eBioscience, 1:100), PE-conjugated anti-CD29 (#25029180, eBioscience, 1:100), APC-conjugated anti-CD24 (#562349, BD Biosciences, 1:100), PE-conjugate anti-CD24 (#553262, BD Biosciences, 1:100), PE-conjugated anti-CD61 (#561910, BD Biosciences, 1:100), PerCP-Cy™5.5 Mouse Lineage Antibody Cocktail (#561317, BD Biosciences, 1:100), anti-TET2 (#36449, Cell Signaling Technology, 1:100), and Alexa Fluor® 488-conjugated Anti-rabbit IgG (H + L) F(ab’)2 Fragment (#A-11070, ThermoFisher Scientific, 1:500). Stained cells were subjected to BD Canto II analysis (BD FACS Diva 8), and flow cytometry data (mean% ± SD) was analyzed by FCS express 6 (Denovo Software) from three independent experiments with gating boundaries determined by using antibody isotype controls. FACS sorted cells were fixed and permeabilized, stained with Alexa Fluor® 594-conjugated Cytokeratin 8 Antibody (#NB120-9287AF594, Novus Biologicals, 1:200), Alexa Fluor® 647-conjugated Cytokeratin 14 Antibody (#NBP2-47720AF647, Novus Biologicals, 1:200)19 (link),20 (link), and then subjected to BD FACSAria FACS sorting followed by standard qRT-PCR analysis.
5
Flow Cytometry Characterization of ASCs
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To confirm the ASC-progenitor phenotype, flow cytometry was performed after 3 days following the protocol in the previous subsection. Briefly, after tissue digestion ASCs (8 × 10 4 ) were washed, resuspended in 1% BSA in PBS, and incubated with FITC-conjugated anti-CD29, APC-conjugated anti-CD31, FITC-conjugated anti-CD44, FITCconjugated anti-CD45, and PE-conjugated anti-CD90 (BD Bioscience, San Jose, CA,USA), respectively, for 1 h at 4°C. Corresponding isotype-matched antibodies were used as controls. FACS analysis was performed using an Accuri C6 cytometer (BD, Franklin Lakes, NJ, USA).
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