Whole pancreata were removed from WT and β14-3-3ζ–KO mice, weighed, fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned to 6 μm thickness. A minimum of 3 sections, 72 μm apart, were used in all studies. Antigen retrieval at 95°C was performed using EZ-Retriever System (Biogenex) with 10 mM sodium citrate buffer. β Cell proliferation was assessed, as described above. The In Situ Cell Death Detection Kit (TUNEL; Roche Applied Sciences) was used to measure β cell apoptosis (Roche; ref. 15 (link)). To measure β cell mass in pancreatic sections, IHC was performed with an insulin antibody (1:200 dilution; Ab 3014, Cell Signaling Technology) and the SignalStain DAB Substrate Kit (Cell Signaling Technology). Hematoxylin was used for counterstaining. Slides were monitored using a high-resolution scanner (Aperio ImageScope 12.3.3) to assess the areas of insulin+ β cells and the whole pancreas, followed by calculating β cell mass (total β cell area/total pancreas weight).
Ab 3014
Manufactured by Cell Signaling Technology
Ab 3014 is a primary antibody product offered by Cell Signaling Technology. It is a highly specific and well-characterized antibody that recognizes a target protein. The core function of Ab 3014 is to serve as a detection tool for the target protein in various research and analytical applications.
Automatically generated - may contain errors
2 protocols using ab 3014
1
Quantifying Pancreatic Beta Cell Mass
2
Quantifying β-Cell Mass and Apoptosis
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Whole pancreata were removed from wild type (WT) and β14-3-3ζKO mice, weighed, fixed in 4% paraformaldehyde, embedded in paraffin and sectioned to 6-μm thickness. A minimum of 3 sections, 72 μm apart, were used in all studies. Antigen retrieval at 95ºC was performed using EZ-Retriever® System (Biogenex, Fremont, CA) with 10mM sodium citrate buffer. β-cell proliferation was assessed, as described above. The In Situ Cell Death Detection Kit (TUNEL; Roche Applied Sciences, Penzberg, Germany) was used to measure β-cell apoptosis (Roche) (Lim et al., 2016) . To measure β-cell mass in pancreatic sections, immunohistochemistry was performed with an insulin antibody (1:200 dilution; Ab 3014, Cell Signaling Technology, Danvers, MA) and the SignalStain® DAB Substrate Kit (Cell Signaling Technology).
Hematoxylin was used for counter-staining. Slides were monitored using a high-resolution scanner (Aperio ImageScope 12.3.3) to assess the areas of insulin-positive β-cells and the whole pancreas, followed by calculating β-cell mass (total b-cell area / total pancreas weight).
Hematoxylin was used for counter-staining. Slides were monitored using a high-resolution scanner (Aperio ImageScope 12.3.3) to assess the areas of insulin-positive β-cells and the whole pancreas, followed by calculating β-cell mass (total b-cell area / total pancreas weight).
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