Staining was performed as previously reported (39 (link)). In brief, tissue sections were cut at 4μm and deparaffinized in xylene and absolute alcohol. Endogenous peroxidase activities were blocked with a solution of 3% hydrogen peroxide and absolute alcohol 1:1 for 15 minutes. Heat induced antigen retrieval was performed with a pressure cooker (122+/−2°C) for 45 seconds at 15+/−5 PSI in citrate buffer (pH6.0). Immunohistochemistry was performed using a polyclonal rabbit antibody to FOXJ1 (HPA005714, Sigma-Aldrich) at a dilution of 1:75 incubated for 45 minutes at room temperature followed by labeled polymer-HRP anti-rabbit IgG incubated 30 minutes and visualized with 3,3′-Diaminobenzidine (DAB) (brown in color: Dako Envision System). The slides were counterstained with hematoxylin. Images were acquired using an Olympus BX41 microscope and an Olympus DP26 digital camera.
Hpa005714
Manufactured by Merck Group
The HPA005714 is a laboratory equipment product manufactured by Merck Group. It is a device designed for use in research and scientific applications. The core function of this product is to facilitate various laboratory tasks and experiments, but a detailed description cannot be provided while maintaining an unbiased and factual approach.
Automatically generated - may contain errors
Lab products found in correlation
Dp26 digital camera, by Olympus (1 mentions)
3 3 diaminobenzidine (dab), by Agilent Technologies (1 mentions)
Bx41 microscope, by Olympus (1 mentions)
X0936, by Agilent Technologies (1 mentions)
Rnase inhibitor, by Promega (1 mentions)
Mab1378, by Merck Group (1 mentions)
Itga6, by Merck Group (1 mentions)
Donkey anti rat igg cy3, by Jackson ImmunoResearch (1 mentions)
Cy3 donkey anti rabbit igg, by Jackson ImmunoResearch (1 mentions)
Obt0030, by Bio-Rad (1 mentions)
Donkey anti mouse igg alexa 488, by Jackson ImmunoResearch (1 mentions)
Bz x710, by Keyence (1 mentions)
4 protocols using hpa005714
1
Immunohistochemical Staining of FOXJ1 in Tissue Sections
2
Immunohistochemical Staining for FOXJ1 in Ependymoma
3
Immunophenotyping of Trypsinized Cells
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Cells were harvested by Trypsin and washed with PBS once before fixation with 4% PFA. After blocking with block buffer (IF buffer with 1% BSA) for 30 min, cells were stained with FOXJ1 (RRID: AB_1078902; HPA005714; Sigma-Aldrich) and ITGA6 (RRID: AB_2128317; MAB1378; Millipore) antibodies for 30 min, followed by incubation with secondary antibodies (Alexa Fluor 488 anti-rat and 647 anti-rabbit). The cells were suspended in 2% FBS and analyzed by flow cytometry or cell sorting. When RNA was required to be extracted and recovered after cell sorting, RNase inhibitors (N2611; Promega) were included in all of the above buffers at a concentration of at least 1:100.
4
Immunofluorescence Staining of Cilia and Stem Cell Markers
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Cells were fixed in 4% paraformaldehyde for 6 min at room temperature. The cells were then processed for immunofluorescence using mouse antibody against β-Tubulin IV (1:200, Sigma-Aldrich, T7941) to stain cilia. Rabbit anti-FoxJ1 polyclonal antibody (1:200, Sigma, HPA005714), mouse anti-nestin monoclonal antibody (1:400, BD, BD611658), and rat anti-BrdU monoclonal antibody (1:200, AbD serotec, OBT0030) were also used as primary antibodies. Donkey anti-mouse IgG-Alexa488 (1:500, Jackson ImmunoResearch, 715-545-150), donkey anti-rabbit IgG-Cy3 (1:500, Jackson ImmunoResearch, 712-165-153), and donkey anti-rat IgG-Cy3 (1:500, Jackson ImmunoResearch, 711-165-152) were used as secondary antibodies. Nuclei were counterstained with DAPI. Fluorescent images of cells were obtained using a fluorescent microscope (BZ-X710, Keyence). The total number of cells was determined by counting DAPI stained nuclear signals using Fiji (ImageJ) (Schindelin et al. 2012) . The number of FoxJ1 positive cells was also determined using Fiji. Numbers of BrdU or nestin-positive cells or multi-ciliated cells were counted manually.
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