SVZ cells were exposed to BrdU (10 μM, Sigma-Aldrich Co. LLC) 4 h before the end of the experiment (48 h post-transfection). Thereafter, cells were fixed in 4% PFA. Fixed cells were permeabilized with 1% Triton X-100 for 30 min at RT and BrdU was exposed by incubating the fixed cells with 1 M HCL for 40 min at 37 °C. Nonspecific binding sites were blocked with 6% bovine serum albumin (BSA) in PBS containing 0.3% Triton X-100 for 1 h, followed by incubation with anti BrdU Alexa-Fluor 594 conjugated antibody (Life Technologies) prepared in PBS containing 0.3% BSA and 0.3% Triton X-100 for 2 h at RT. Cells were then stained with Hoechst-33342 and mounted in Fluoroshield Mounting Medium (Abcam Plc.). Photomicrographs of BrdU incorporation were taken using a confocal microscope (AxioObserver LSM 710, Carl Zeiss).
Anti brdu alexa fluor 594 conjugated antibody
Manufactured by Thermo Fisher Scientific
The Anti BrdU Alexa-Fluor 594 conjugated antibody is a laboratory reagent used for the detection and quantification of bromodeoxyuridine (BrdU) incorporation into cellular DNA. The antibody is conjugated with the Alexa Fluor 594 fluorescent dye, allowing for visualization and analysis of BrdU-labeled cells using fluorescence-based techniques.
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2 protocols using anti brdu alexa fluor 594 conjugated antibody
1
BrdU Incorporation in SVZ Cells
2
Quantifying Cell Proliferation with BrdU Labeling
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5-bromo-2'-deoxyuridine (BrdU; 10µM, Sigma-Aldrich Co. LLC) was added to cultures 4 h before the end of the experiment. Thereafter, cells were fixed in 4% PFA and BrdU was exposed following permeabilization with 1% Triton X-100 for 30 min at RT and an incubation with 1 M HCL for 40 min at 37 ºC. Nonspecific binding sites were blocked with 6% BSA and 0.3% Triton X-100 for 1 h, followed by incubation with anti-BrdU Alexa-Fluor 594 conjugated antibody (Life Technologies) prepared in 0.3% BSA and 0.3% Triton X-100 for 2 h at RT. Cells where then stained with Hoechst-33342 and mounted in Fluoroshield Mounting Medium (Abcam Plc.). Photomicrographs of BrdU incorporation were taken using a confocal microscope (AxioObserver LSM 710, Carl Zeiss).
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