Sk 4805

Manufactured by Vector Laboratories

SK-4805 is a piece of laboratory equipment manufactured by Vector Laboratories. It is designed to perform a core function within the lab environment. The detailed specifications and intended use of this product are not available at this time.

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3 protocols using sk 4805

Whole-mount in situ hybridization and Blimp1 immunohistochemistry

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Whole-mount in situ hybridisation analysis was performed as before^{50 (link)}, using antisense riboprobes for Bmp2^{64 (link)}, Bmp4^{13 (link)} and Gdf3^{28 (link)}. For Blimp1 immunohistochemistry, E7.5 decidua were fixed overnight in 4% PFA, dehydrated in ethanol, embedded in paraffin wax and sectioned (6 μm). Dewaxed sections were subjected to antigen retrieval by boiling for 1 h in Tris/EDTA (pH 9.0) and permeabilized for 10 min in 0.1% Triton X-100 in TBS. Sections were subsequently blocked with 10% normal goat serum in TBS. Sections were incubated in rat monoclonal anti-Blimp1 (1:500 dilution, sc-130917, Santa Cruz Biotechnology) in block overnight at 4 °C and signal-amplified with rabbit anti-rat secondary antibody (AI-4001, Vector Laboratories) for 45 min at RT followed by peroxidase blocking for 20 min at RT and development with Envison System-HRP for rabbit antibodies (K4011, DAKO) and Vector Red substrate (SK-4805, Vector Laboratories). Sections were lightly counter stained with haematoxylin, coverslipped and imaged.
Haematoxylin and eosin staining was performed as previously described^{36 (link)}.

Senft A.D., Bikoff E.K., Robertson E.J, & Costello I. (2019). Genetic dissection of Nodal and Bmp signalling requirements during primordial germ cell development in mouse. Nature Communications, 10, 1089.

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Immunohistochemical Detection of SARS-CoV-2 in Formalin-Fixed Tissues

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Formalin-fixed and paraffin-embedded patient tissue samples were obtained using the University of Calgary Research Ethics Board protocol REB14-1965. Hematoxylin and eosin staining was carried out as per standard protocol by the Alberta Precisions Laboratories (Calgary, Canada). Immunohistochemistry was performed on the slides as described.10 (link) In brief, endogenous peroxidase activity was inhibited with 3% hydrogen peroxide. The slides were then blocked with 4% goat serum before being incubated with a primary mouse antibody against SARS-CoV-2 (GeneTex GTX632604, Clone1A9) overnight at 4°C. After washing with phosphate buffer solution (5 times, 5 min each), the slides were incubated with biotinylated goat anti-mouse IgG secondary antibody overnight (BA-9200-1.5, Vector Laboratories). Chromogen readout was achieved using the avidin/biotin peroxidase system (PK-6100, Vector Laboratories) and NovaRed substrate (SK-4805, Vector Laboratories).

Swain L.A., Ambasta A., Munhoz E.P., Omodon O., Urbanski S.J, & Nguyen H.H. (2023). Acute severe hepatitis as a presenting symptom in clinically stable patients admitted with SARS-CoV-2 Omicron infection. Hepatology Communications, 7(4), e0115.

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Immunohistochemical Analysis of DCLK1+ Cells

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Unstained 5-micron sections derived from the formalin-fixed and paraffin-embedded blocks were deparaffinized and hydrated by routine procedures. Sodium citrate buffer (pH 6.0) was used as the antigen retrieval. The primary antibodies at diluted concentrations were incubated overnight at room temperature. The primary antibodies are listed in Supplementary Table 1. For immunohistochemistry (IHC), the secondary antibodies used were Dako LSAB+system-HRP (universal) and Envision+system-HRP (anti-rabbit and anti-mouse polymer-HRP). For double IHC, anti-rabbit and anti-mouse polymer-AP kits were purchased from Vector (MP-5401 and MP5402), including the substrate kits for red peroxidase (SK-4805), red alkaline phosphatase (SK-5100) and blue alkaline phosphatase (SK-5300). For immunofluorescence (IF) assay, the fluorophore-conjugated secondary antibodies (Jackson ImmunoResearch) were incubated at room temperature for 2 hours. All other procedures were done according to the manufactures’ instructions. The percentage of DCLK1+ cells for each experimental group (ADM, PanIN, IPMN, and normal) was determined by counting DCK1+ cells and total ductal epithelial cells present in the pancreata of ten randomly chosen mice within each group (MT-TGF-α, KP, AKP GEMM, and Cre-negative control), and at least three different sections of each individual pancreas were examined.

Qiu W., Remotti H.E., Tang S.M., Wang E., Dobberteen L., Youssof A.L., Lee J.H., Cheung E.C, & Su G.H. (2018). Pancreatic DCLK1+ Cells Originate Distinctly from PDX1+ Progenitors and Contribute to the Initiation of Intraductal Papillary Mucinous Neoplasm in Mice. Cancer letters, 423, 71-79.

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