Primary human bladder epithelial cells were seeded on 22×22 mm glass coverslips and were cultured with 10uM 5-Bromo-2’-deoxyuridine (Sigma) and different concentrations of various growth factors as indicated in the figure legends for 24 hours in Keratinocyte SFM medium. The next day, the cells were fixed in 4% paraformaldehyde and permeabilized in blocking buffer (0.1% saponin, 1% fish gelatin, 5% normal mouse serum in PBS) for 30 min in room temperature. Then the samples were incubated in 1M HCL for 1 hour at room temperature, followed by three times PBS wash and another 30 min incubation with blocking buffer. The cells were then incubated with FITC conjugated Mouse Anti-Human BrdU antibody with DNase (1:100 dilution in blocking buffer) overnight in 4°C. Then coverslips were mounted with Fluoroshield with DAPI reagent (Sigma) and examined using a Leica SP5 confocal microscope. In ImageJ, the cells with positive staining of BrdU were counted.
Fluoroshield with dapi reagent
Manufactured by Merck Group
Fluoroshield with DAPI reagent is a laboratory product that contains the fluorescent dye DAPI (4',6-diamidino-2-phenylindole). DAPI is a DNA-binding agent that emits blue fluorescence when bound to double-stranded DNA. This product is used for the detection and visualization of DNA in various applications, such as fluorescence microscopy and flow cytometry.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using fluoroshield with dapi reagent
1
Quantifying Proliferation in Bladder Cells
2
Quantifying Proliferation in Bladder Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Primary human bladder epithelial cells were seeded on 22×22 mm glass coverslips and were cultured with 10uM 5-Bromo-2’-deoxyuridine (Sigma) and different concentrations of various growth factors as indicated in the figure legends for 24 hours in Keratinocyte SFM medium. The next day, the cells were fixed in 4% paraformaldehyde and permeabilized in blocking buffer (0.1% saponin, 1% fish gelatin, 5% normal mouse serum in PBS) for 30 min in room temperature. Then the samples were incubated in 1M HCL for 1 hour at room temperature, followed by three times PBS wash and another 30 min incubation with blocking buffer. The cells were then incubated with FITC conjugated Mouse Anti-Human BrdU antibody with DNase (1:100 dilution in blocking buffer) overnight in 4°C. Then coverslips were mounted with Fluoroshield with DAPI reagent (Sigma) and examined using a Leica SP5 confocal microscope. In ImageJ, the cells with positive staining of BrdU were counted.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!