Following perfusion, as described in animals and operative procedure, the skin from the back was removed and connective tissue covering the CMM was carefully dissected off of the whole back while leaving the skin intact. The back was then cut in half along the midline and one side was randomly selected for staining. The back half was permeabilized for 1.5 hours in a solution of 1% Triton X and 5% DMSO (Sigma-Aldrich) in 1× PBS and then stained with α-bungarotoxin conjugated to Alexa Fluor 594 (Life Technologies) to identify the postsynaptic NMJs. The back halves were then whole mounted onto slides using ProLong Gold Antifade (Life Technologies). CMM tissue from WT mice were also blocked in the same permeabilization solution with 5% goat serum (Jackson ImmunoResearch Laboratories Inc., West Grove, PA). We blocked the tissue with Mouse on Mouse blocking reagent then stained the whole CMM from WT mice for synaptophysin (SV2 antibody from DSHB) and Schwann cells (S-100 antibody from Sigma-Aldrich) to identify endogenously expressed markers of the neuromuscular synapse.
S 100 antibody
Manufactured by Merck Group
Sourced in United States
The S-100 antibody is a laboratory research tool used for the detection and identification of S-100 protein, a calcium-binding protein found in certain cell types. The S-100 antibody is commonly used in immunohistochemistry and Western blotting techniques to study the distribution and expression of S-100 protein in various tissues and cell samples.
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Lab products found in correlation
2 protocols using s 100 antibody
1
Neuromuscular Junction Visualization in Mouse Skeletal Muscle
2
Immunohistochemical Analysis of Nerve Grafts
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Grafts were fixed in 10% v/v neutral buffer formalin solution (Sigma-Aldrich, Darmstadt, Germany) paraffin embedded and sectioned. Then, the slides were deparafinnized, rehydrated and blocked. Dako Envision Flex kit was used for the immunohistochemistry assay according to manufacturer’s instructions (Dako, Agilent, Glostrup, Denmark). Briefly, nerve graft sections were incubated at 4 °C over night with rabbit anti-neurofilament 200 (nf 200) antibody (1:80, Sigma, St. Louis, MO, USA) to identify axons and S100 antibody (1:100, Sigma, St. Louis, MO, USA) to identify Schwann cells. Briefly, washes were performed, and addition of horseradish peroxidase (HRP) conjugated with goat secondary antibody against rabbit and mouse was performed. The slides were incubated at Room Temperature (RT) for 45 min. Finally, 3′3 diaminobenzidine (DAB) was added to the slides. Slides were visualized by light microscopy and images were acquired with ΙC Capture 2.2 software and processed with imageJ software version 1.52g.
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