The bromodeoxyuridine (BrdU) incorporation assay was used to identify proliferating cells. Cells seeded on gels and TCP were incubated with medium containing BrdU labeling agent (dilution 1 : 200; Abcam) for 24 hours. Cells were then fixed with 4% paraformaldehyde, denatured by 2 M HCl for 10 minutes, permeabilized with 0.5% Triton-X-100 (Sigma Aldrich) for 30 minutes and blocked with 1% bovine serum albumin. Anti-BrdU antibody (dilution 1 : 200, Thermo-scientific, USA) was added to the BrdU labeled cells and was incubated overnight at 4 °C. After several washes with PBS, secondary antibody Alexa Fluor®568 Rabbit anti-mouse (dilution 1 : 1000, Thermo-scientific, USA) was added along with a nuclear counterstain (Hoechst, 33 342, Thermo-scientific, USA). Labeled cells were imaged using EVOS FL Auto (Life Technologies, USA) at 10× magnification.
Alexa fluor 568 rabbit anti mouse
Manufactured by Thermo Fisher Scientific
Sourced in United States
Alexa Fluor®568 Rabbit anti-mouse is a fluorescently labeled secondary antibody. It is designed to detect and visualize mouse primary antibodies in various immunochemical applications.
Automatically generated - may contain errors
2 protocols using alexa fluor 568 rabbit anti mouse
1
Quantifying Cell Proliferation with BrdU Assay
2
BrdU Incorporation Assay for Proliferating Cells
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The bromodeoxyuridine (BrdU) incorporation assay was used to identify proliferating cells. Cells seeded on gels and TCP were incubated with medium containing BrdU labeling agent (dilution 1 : 200; Abcam) for 24 hours. Cells were then fixed with 4% paraformaldehyde, denatured by 2 M HCl for 10 minutes, permeabilized with 0.5% Triton-X-100 (Sigma Aldrich) for 30 minutes and blocked with 1% bovine serum albumin. Anti-BrdU antibody (dilution 1 : 200, Thermo-scientific, USA) was added to the BrdU labeled cells and was incubated overnight at 4 °C. After several washes with PBS, secondary antibody Alexa Fluor®568 Rabbit anti-mouse (dilution 1 : 1000, Thermo-scientific, USA) was added along with a nuclear counterstain (Hoechst, 33 342, Thermo-scientific, USA). Labeled cells were imaged using EVOS FL Auto (Life Technologies, USA) at 10× magnification.
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