Cases and controls brain samples were derived from the Mount Sinai Neuropathology Brain Bank. Inclusion criteria were individuals with a neuropathological diagnosis of Parkinson’s disease for cases and cognitively normal with no or only age-related neuropathological changes for controls. Formalin-fixed paraffin-embedded (FFPE) sections (5μm) were prepared from substantia nigra blocks, mounted on positively charged slides, and baked overnight at 70 °C. Immunohistochemistry (IHC) was performed on a Ventana Benchmark XT automatic staining platform (Roche Diagnostics, Indianapolis, IN) according to the manufacturer’s protocol with reagents and antibodies acquired from the same lot. Antigen retrieval was done using CC1 buffer (Tris/Borate/EDTA buffer, pH 8.0–8.5, Roche Diagnostics) for 1 h followed by primary antibody incubation. All primary antibodies were diluted in antibody dilution buffer (ABD24, Roche Diagnostics). Primary antibodies were incubated for 36 min (mOXDJ1, 1:400, Abcam) or 28 min (GFAP, 1:10, Ventana, 760-4345) followed by either 3,3’-diaminobenzidine (DAB) or alkaline phosphatase for visualization. For slides that were double-labeled, both DAB and alkaline phosphatase were used for visualization. All slides were counterstained with hematoxylin and coverslipped.
Abd24
Manufactured by Roche
The ABD24 is an automated blood drawing device designed for clinical laboratory use. It is capable of collecting blood samples from patients in a consistent and efficient manner. The device handles the blood collection process, including skin disinfection, vein detection, and sample transfer to appropriate containers. The core function of the ABD24 is to automate the blood drawing procedure to improve reliability and reduce human error.
Automatically generated - may contain errors
2 protocols using abd24
1
Parkinson's Disease Brain Immunohistochemistry
2
Immunohistochemistry Staining of Organoids
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Organoids were washed with PBS, fixed in fresh 4% PFA at RT for 30-60 min (depending on organoid size) on a shaker and stored in PBS at 4°C until embedding. The samples were embedded in paraffin and 5 μm sections were prepared and mounted on positively charged slides. The sections were baked O/N at 70°C. IHC staining was performed on a Ventana Benchmark XT automatic staining platform (Roche Diagnostics #06468373001) according to the manufacturer's guidelines. Antigen retrieval was performed using the CC1 buffer (Tris/Borate/EDTA buffer, pH 8.0 - 8.5; Roche Diagnostics #950-224) for 1 hour. The samples were incubated in primary antibodies diluted in antibody dilution buffer (ABD24; Roche Diagnostics #05280524001) and the incubation time was optimized for each antibody. The slides were then incubated in either 3,3'-diaminobenzidine (DAB) or alkaline phosphatase for visualization, and both reagents were used for slides that were double-stained. Hematoxylin was used for nuclear counterstaining, and the slides were coverslippped. Table S4 (bottom) and the key resources table list the antibodies and dilutions used for IHC staining in this study.
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