Cell tissue staining kit

Manufactured by R&D Systems

Sourced in United States

The Cell & Tissue Staining Kit is a comprehensive set of reagents designed for the visualization and analysis of cellular and tissue components. The kit provides the necessary tools for staining a variety of target structures, enabling researchers to study the morphology, distribution, and expression patterns of specific cellular and extracellular elements.

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Lab products found in correlation

Bx51 microscope, by Nikon (1 mentions) Ab34710, by Abcam (1 mentions) Ab7778, by Abcam (1 mentions)

5 protocols using cell tissue staining kit

Quantifying Neointimal Formation and Macrophage Infiltration in Balloon-Injured Arteries

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Neointimal formation in balloon-injured arteries was evaluated 14 days after injury. Carotid arteries from experimental groups were embedded in paraffin and 5 µm cross sections were prepared. Morphometric analysis via computerized image analysis system (ImageJ v1.43) was performed in vessel sections stained with hematoxylin-eosin (H-E). The average of the nine sections was used as the value for one animal. The parameter for analysis of neointimal formation was neointimal to medial area ratio (N/M)^{41 (link),54 (link)}. Macrophage infiltration was evaluated on day 14 after injury using primary antibodies for CD68. In brief, the formalin-fixed/paraffin-embedded sections were deparaffinized, followed by epitope masking with Antigen Retroviral Reagent (R&D Systems, catalog number CTS016). The sections were then treated with Serum Blocking Reagent (R&D Systems, catalog number 865015), and incubated with CD68 antibody for overnight at 4 °C. The CD68 staining was assessed using a HRP-DAB cell staining kit (R&D Systems, Cell & Tissue Staining Kit) according to the protocol provided by the manufacturer. Sections were counterstained with hematoxylin, mounted, and photographed using a Leica microscope.

Sorrentino S., Iaconetti C., De Rosa S., Polimeni A., Sabatino J., Gareri C., Passafaro F., Mancuso T., Tammè L., Mignogna C., Camastra C., Esposito G., Curcio A., Torella D, & Indolfi C. (2018). Hindlimb Ischemia Impairs Endothelial Recovery and Increases Neointimal Proliferation in the Carotid Artery. Scientific Reports, 8, 761.

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Immunohistochemical Detection of CD215 in Vitiligo

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Immunohistochemical studies for CD215 were performed on 5-μm sections from formalin-fixed, paraffin-embedded ellipse biopsy specimens from vitiligo patients using a Cell & Tissue staining kit according to the manufacturer’s instructions (R&D Systems) with a slight modification. Briefly, after deparaffinization and rehydration, the antigen retrieval was performed with citrate buffer (pH 6.0). The sections were permeabilized with PBST (0.2% Triton X-100 in PBS) and then blocked with blocking reagents of peroxidase, serum, avidin, and biotin sequentially. The sections were blocked with Human TruStain (BioLegend) and incubated with goat anti-human CD215 (6 to 10 μg/ml; sc-1524 or AF-247) or goat immunoglobulin G isotype control (R&D Systems) overnight at 4°C, followed by incubation with biotinylated secondary antibody at room temperature for 1 hour. The staining was visualized using the R&D Systems HRP-DAB detection reagent. All the sections were counterstained with hematoxylin, and images were taken using an Olympus BX51 microscope with Nikon NIS Elements software version 3.10.

Richmond J.M., Strassner J.P., Zapata L J.r., Garg M., Riding R.L., Refat M.A., Fan X., Azzolino V., Tovar-Garza A., Tsurushita N., Pandya A.G., Tso J.Y, & Harris J.E. (2018). Antibody blockade of IL-15 signaling has the potential to durably reverse vitiligo. Science translational medicine, 10(450), eaam7710.

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Immunohistochemical Analysis of Tumor Angiogenesis

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Tumor tissues were fixed in 4% paraformaldehyde for 12 h. The samples were incubated in 30% sucrose for 12 h, and then embedded in OCT compound and cut to 5 µm sections. The slides were incubated with specific antibodies for anti-CD31 (1:200, cat. no. 553370, BD Biosciences) and anti-PTPN12 (1:100, cat. no. sc-271351, Santa Cruz Biotechnology). followed by HRP-conjugated anti-mouse or anti-rabbit IgG using the cell & tissue staining kit (R&D systems). The reaction was then developed with DAB substrate.

Kim O., Hwangbo C., Tran P.T, & Lee J.H. (2022). Syntenin-1-mediated small extracellular vesicles promotes cell growth, migration, and angiogenesis by increasing onco-miRNAs secretion in lung cancer cells. Cell Death & Disease, 13(2), 122.

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Histological and Immunohistochemical Analysis of Intervertebral Disc Degeneration

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The specimens were washed in PBS, fixed with 4% formaldehyde in PBS overnight, decalcified in 10% formic acid solution, dehydrated through a graded series of ethanol, embedded in paraffin, and sectioned at a thickness of 5 μm. For histological analysis, sections were deparaffinized, rehydrated, and stained with hematoxylin and eosin (HE), Safranin-O (SO) and Masson Trichrome. For immunohistochemical (IHC) staining, rehydrated sections were pre-treated with papain solution for 15 min, incubated with NR4A1 antibody (sc-7014, Santa Cruz Inc, USA) at 1:100 dilutions (2μg/ml) for 1 hour and detected by a cell & tissue staining kit (R&D systems Inc., Minneapolis, MN, USA) according to the manual. All sections were counterstained with hematoxylin. Positive and negative controls were also stained with the study samples to validate the immunohistochemical staining.
To quantify the histological results, we calculated the histological scores based on the method of Masuda et al [54 (link)]. Briefly, the slides were graded based on the histological appearance of 4 parameters: the annulus fibrosus, the border between the annulus fibrosus and NP, the cellularity of NP, and the matrix of NP. Each of the 4 parameters was given a grade of 1, 2, or 3. The sum of the grades for each parameter yielded a total grade. Total grades ranged from 4 to 12 with 12 representing severe degeneration.

Feng G., Zhang Z., Dang M., Zhang X., Doleyres Y., Song Y., Chen D, & Ma P.X. (2017). Injectable nanofibrous spongy microspheres for NR4A1 plasmid DNA transfection to reverse fibrotic degeneration and support disc regeneration. Biomaterials, 131, 86-97.

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Immunohistochemical Analysis of Collagen I and III

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An avidin-biotin-peroxidase complex commercial method (cell & tissue staining kit, R&D Systems, Inc., USA) was used for IHC. Briefly, 4-mm-thick paraffin wax sections were mounted on slides, which were dried for 30 min in an oven (60–70 centigrade degree) and deparaffinized in xylene. The slides were then placed in changes of ethanol for 2 mins each. Washing in buffer solution was performed between steps. The slides were then placed in 3% hydrogenperoxide for 15 mins and were subsequently incubated in avidin block for 15 mins, biotin block for 15 mins, primary antibody (Anti-Collagen I antibody, Abcam, USA. ab34710; Anti-Collagen III antibody, Abcam, USA, ab7778) for 12 h at 4 centigrade degree, and biotinylated secondary antibody for 1 hour. The reagent incubation was performed with streptavidin peroxidase for 15 mins. A 1 min Mayer’s hematoxylin counter stain was used. The slides were dehydrated, cleared with xylene, and mounted with permanent mounting medium. Finally pictures were analyzed by IPP 6.0 software.

Li C., Wang Y., Qiu Q., Shi T., Wu Y., Han J., Chai X, & Wang W. (2014). Qishenyiqi Protects Ligation-Induced Left Ventricular Remodeling by Attenuating Inflammation and Fibrosis via STAT3 and NF-κB Signaling Pathway. PLoS ONE, 9(8), e104255.

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