Briefly, 30 μm sections of the formalin-fixed brain were sectioned on a vibratome and collected in PBS containing 0.01% sodium azide (Sigma-Aldrich). Free-floating tissues were treated for antigen retrieval (Abcam, Cambridge, UK) in the microwave by heating for 20 min. The tissues were permeabilized in 0.25% Triton X-100 for 1 h, followed by blocking with 10% normal goat serum for 1 h at room temperature. Subsequently, tissue sections were incubated with antibodies against the glial fibrillary acidic protein (GFAP, 1:400, Sigma-Aldrich) and aquaporin-4 (AQP-4, 1:500, Abcam) overnight at 4 °C. The tissues were incubated with Alexa Fluor 488-labeled goat anti-mouse-IgG (1:1000; #A32723, Invitrogen, Carlsbad, CA, USA) or Alexa Fluor 633-labeled goat anti-rabbit-IgG (1:1000; #A32731, Invitrogen), correspondingly, for 1 h at room temperature. The slides were mounted with fluorescence mounting medium (Dako).
Antigen retrieval
Manufactured by Abcam
Sourced in United Kingdom
Antigen retrieval is a laboratory technique used to expose antigenic sites within a sample, typically tissue or cells, prior to immunodetection. It enhances the accessibility of target antigens to antibodies, improving the sensitivity and specificity of immunohistochemical or immunocytochemical analyses.
Automatically generated - may contain errors
Lab products found in correlation
Fluorescence mounting medium, by Agilent Technologies (1 mentions)
Glial fibrillary acidic protein (gfap), by Merck Group (1 mentions)
Alexa fluor 488 labeled goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Sodium azide, by Merck Group (1 mentions)
Confocal sp8 smd microscope, by Leica (1 mentions)
Click it edu imaging kit, by Thermo Fisher Scientific (1 mentions)
Vectashield mounting media, by Vector Laboratories (1 mentions)
Antigenfix, by Diapath (1 mentions)
2 protocols using antigen retrieval
1
Immunohistochemical Staining of Brain Tissue
2
Measuring Pancreatic Beta Cell Proliferation
Check if the same lab product or an alternative is used in the 5 most similar protocols
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EdU was intraperitoneal injected (100 μg/g) into mice. The pancreas was harvested 1 h later, fixed in Antigenfix (Diapath) and embedded in paraffin blocks. Sections (5-μm-thick) were cut, deparaffinized in xylene, rehydrated followed by antigen retrieval (Abcam). EdU staining was conducted using the Click-iT™ EdU imaging kit (Invitrogen) according to the manufacturer’s protocol. Anti-insulin was used to identify β-cells. Slides were mounted with Vectashield mounting media (Vector Laboratories Inc). For each mouse eight randomly selected sections were included for analysis. Images were captured using a Leica SP8 SMD confocal microscope and EdU-positive β-cells were counted.
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