After the rabbits were sacrificed, the knee joint samples were removed and fixed in 10% neutral buffered formalin for 3 days and were then dehydrated through an alcohol gradient (30%–100%) and embedded in paraffin wax. The specimens were cut to a thickness of 5 μm along the longitudinal axis of the tendon. These sections were treated with haematoxylin and eosin (H&E) and Masson’s trichrome stain (Masson’s) for histological evaluation. The sections stained with H&E and Masson’s were observed using a light microscope (Eclipse 80i, Nikon), and the images were captured using a camera (DS-5M, Nikon). For immunohistochemistry staining, endogenous peroxidase was blocked by incubation with 3% (v/v) hydrogen peroxide in methanol for 10 mins. Then, the sections were blocked for 20 mins with a blocking reagent containing goat serum in PBS after washing three times with PBS. After overnight incubation at 4°C with a primary antibody, the sections were washed and then incubated with a secondary antibody (MaxVision kit; Maixin Biotechnology) for 15 mins at room temperature. Dimethyl aminoazobenzene (DAB) (Simple Stain DAB Solution, Maixin Biotechnology) was used for colour development. Hematoxylin staining was used to reveal the nuclei. The results were evaluated by three individuals who were blinded to the treatments.
Simple stain dab solution
Manufactured by Maixin Group
Sourced in China
The Simple Stain DAB Solution is a laboratory reagent used for immunohistochemical (IHC) staining procedures. It is designed to provide a brown chromogenic reaction for the detection of target antigens in tissue samples. The solution contains 3,3'-Diaminobenzidine (DAB), a commonly used chromogen, which reacts with the enzyme label (e.g., horseradish peroxidase) to produce a visible brown staining in the presence of the target antigen.
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Lab products found in correlation
2 protocols using simple stain dab solution
1
Histological and Immunohistochemical Analysis of Rabbit Knee Joints
2
Immunohistochemical Analysis of Collagen I
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The sections of PET artificial ligament in the articular cavity were deparaffinized and rehydrated. Endogenous peroxidase was blocked for 30 min with 0.3% hydrogen peroxide at 37 °C, and further for 20 min with a blocking reagent (PBS containing goat serum). The sections were washed twice with PBS buffer and incubated with the primary antibody (rabbit anti-dog COL1, dilution 1: 50, Bioworld technology co Ltd., Nangjing, China) at 4 °C overnight. After washing, the sections were incubated with a secondary antibody (MaxVision kit, Maixin Biotechnology, Fuzhou, China) for 15 min at room temperature, and then treated with dimethylaminoazobenzene (Simple Stain DAB Solution, Maixin Biotechnology, Fuzhou, China) for 5 min, followed by counter-staining with hematoxylin. A DP Manager (Olympus Optical Co., Tokyo, Japan) was used to obtain the digital images, and Image-Pro Plus 6.0 software (Media Cybernetics Corp., Rockville, USA) was used to measure COL1 expression, which was expressed as the mean area value of COL1-positive staining at the graft site with 200× magnification.
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