The experiments were performed as previously described (An et al., 2022 ; Wang et al., 2020 ; Yang et al., 2022 ). The paraffin sections of colon tissue underwent gradient dehydration and were treated with 1 mM EDTA (pH 8.0) in a microwave oven for 30 min for antigenic repair. The sections were incubated with primary antibodies overnight at 4 °C followed by a General purpose SP Kit (ZSGB-Bio) and DAB immunochromogenic reagent (Gene Tech) according to the manufacturer's instructions. The sections were then stained with hematoxylin (Beyotime Biotech). Signals were imaged with an Aperio VERSA 8 (Leica) multifunctional scanner. Primary antibodies used were anti-OTUD4 (Abcam, ab106368), anti-lysozyme (Abcam, ab108508), and anti-Ki67 (CST, 12202S).
General purpose sp kit
Manufactured by ZSGB-BIO
The General purpose SP Kit is a comprehensive set of laboratory equipment designed for a wide range of general-purpose applications. The kit includes essential items such as pipettes, beakers, test tubes, and other basic glassware necessary for conducting various experiments and procedures in a laboratory setting.
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2 protocols using general purpose sp kit
1
Immunohistochemical Analysis of Colon Tissue
2
Immunohistochemical Characterization of Colon Tissue
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Paraffin sections of colon tissues were dewaxed in xylene followed by hydration in gradient ethanol. Antigen retrieval was performed by heating slides in a microwave for 40 min in sodium citrate buffer (pH 6.0). Then the slides were cooled down naturally to room temperature before being quenched in 3% hydrogen peroxide to block endogenous peroxidase activity. The primary antibody diluted in PBS containing 1% BSA (α-SMA, 1:500; ZO-1, 1:400, Occludin, 1:200) was incubated at 4 °C overnight followed by General purpose SP Kit (ZSGB-Bio) and DAB immunochromogenic reagent (Gene Tech) according to the manufacturer's instructions. Subsequently, the slides were counterstained with hematoxylin (Beyotime Biotech) for 15 s and cover slipped. Images were acquired with Aperio VERSA scanner (Leica). The intensities of DAB staining were measured and quantified with integrated optical density or cell intensity by Image Pro Plus 6 (Media Cybernetics). The following immunohistochemical antibodies were used: α-SMA (Sigma, A2547); ZO-1(Bioss, bs-1329R); Occludin (Abcam, ab216327).
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