Dapi containing mounting media

Manufactured by Thermo Fisher Scientific

Sourced in United States

DAPI-containing mounting media is a laboratory product used to stain and mount biological samples for microscopic analysis. It provides a stable platform for preserving and visualizing cellular structures labeled with fluorescent dyes, including the DNA-binding dye DAPI. The media aids in maintaining sample integrity during the mounting process.

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Lab products found in correlation

Lsm 710 confocal microscope, by Zeiss (6 mentions) Lsm 780, by Zeiss (2 mentions) Acti stain 670 phalloidin, by Cytoskeleton (2 mentions) Ix70 microscope, by Olympus (1 mentions) Nb100 60554, by Novus Biologicals (1 mentions) Lamp1, by Cell Signaling Technology (1 mentions) Lsm 510 confocal microscope, by Zeiss (1 mentions) Ab197730, by Abcam (1 mentions) Alexa 488, by Abcam (1 mentions) Fluorescence microscope, by Olympus (1 mentions) Forskolin, by Cell Signaling Technology (1 mentions) Fluorescence microscope, by Zeiss (1 mentions) Anti e cadherin, by Cell Signaling Technology (1 mentions) Ab2907, by Abcam (1 mentions) Anti rabbit igg atto 488, by Merck Group (1 mentions) Anti g quadruplex dna 1h6, by Merck Group (1 mentions) Hela cell, by American Type Culture Collection (1 mentions) Anti mouse igg atto 488, by Merck Group (1 mentions) Iscove s modified dulbecco s medium, by Corning (1 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)

11 protocols using dapi containing mounting media

Immunocytochemical Analysis of Neuronal Cultures

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Primary neuronal cultures grown on cover glass in 12 well tissue culture plates were fixed at 12 days in vitro (DIV12) with 4 % paraformaldehyde for 30 min. Cultures were washed and permeabilized with 0.1 % Triton X-100 and 5 % donkey serum for 1 h at room temperature. Cells on coverglass were then incubated with rabbit anti-APP (C66) and mouse anti-Syt-1 antibodies at 4 °C for overnight. Following washing, cells were again incubated with Alexa Fluor488-conjugated anti-rabbit and Alexa568-conjugated anti-mouse secondary antibodies for 2 h at room temperature. Cells on coverglass were then washed and then mounted on glass slides with the help of DAPI containing mounting media (Life Technologies, Grand Island, NY). Images were acquired on Olympus IX-70 microscope at similar exposure settings and later processed by IP lab software.

Gautam V., D’Avanzo C., Berezovska O., Tanzi R.E, & Kovacs D.M. (2015). Synaptotagmins interact with APP and promote Aβ generation. Molecular Neurodegeneration, 10, 31.

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FISH-based Satellite Heterochromatin Imaging

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SADS assay, which is a fluorescence in situ hybridization (FISH) for detection of peri-centromeric satellite heterochromatin DNA, was performed following the protocol kindly provided by Drs. Sundeep Khosla and Joshua Farr^{76 (link)}. Briefly, paraffin-embedded femoral bone tissue sections were cross-linked with 4% paraformaldehyde and dehydrated in ethanol. The bone tissue sections were then denatured and hybridized in the buffer containing 1.0 μg/mL Cy3-labeled, CENPB-specific (ATTCGTTGGAAACGGGA) peptide nucleic acid FISH probe (Panagene Inc, Korea) for 2 hours at room temperature in the dark. After washing, immunofluorescence staining of the tissue sections was performed using primary antibody against perilipin (Novus Biologicals, NB100-60554, 1:500). Finally, the slides were mounted with DAPI-containing mounting media (Life Technologies). Images were acquired with a Zeiss LSM780 confocal microscope, and the intensity profiles were acquired with Image J software (NIH).

Lemaire V., Koehl M., Le Moal M, & Abrous D.N. (2000). Prenatal stress produces learning deficits associated with an inhibition of neurogenesis in the hippocampus. Proceedings of the National Academy of Sciences of the United States of America, 97(20), 11032-11037.

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Immunofluorescence Analysis of OSMR and LAMP1

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OVCAR4 cells (5 × 10⁴ cells) were treated with control IgG, B14 and B21 (10μg/mL each) and stimulated with OSM (100ng/mL) on coverslip for 16h. Cells were fixed with 4% paraformaldehyde for 10 min and permeabilized with 0.1% triton for 5 min. Cells were then incubated with OSMR (Proteintech) and LAMP1 (Cell Signaling Technology) primary antibodies overnight at 4ºC followed by incubation with goat anti-rabbit Alex Fluor 488 (green) and goat anti-mouse Alexa Fluor 546 secondary antibodies for 1h at room temperature. Cells were mounted with DAPI containing mounting media (Thermo Fisher Scientific) and visualized in Zeiss LSM 510 confocal microscope.

Geethadevi A., Nair A., Parashar D., Ku Z., Xiong W., Deng H., Li Y., George J., McAllister D.M., Sun Y., Kadamberi I.P., Gupta P., Dwinell M.B., Bradley W.H., Rader J.S., Rui H., Schwabe R.F., Zhang N., Pradeep S., An Z, & Chaluvally-Raghavan P. (2021). Oncostatin M Receptor-targeted antibodies suppress STAT3 signaling and inhibit ovarian cancer growth. Cancer research, 81(20), 5336-5352.

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Enrichment and Imaging of CTCs

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CTCs were enriched in 3 mL of peripheral blood samples obtained from two selected lung cancer patients using the aforementioned protocol. The enriched CTC samples were fixed by 4% paraformaldehyde for 20 min at 4 °C. Then the cells were labelled with a CD45 monoclonal antibody conjugated to Alexa 488 (Abcam Cat# ab197730; diluted at 1:100) and a fluorescent conjugates of folic acid (10 nM, Wuxi AppTec, Wuxi, China) for 1 h. After that, sample was washed and mounted with DAPI-containing mounting media (Thermo Fisher Scientific, MA, USA), and subsequently subjected to image analysis using a fluorescence microscope (Olympus, Japan).

Zhou Q., Geng Q., Wang L., Huang J., Liao M., Li Y., Ding Z., Yang S., Zhao H., Shen Q., Pan C., Lou J., Lu S., Chen C, & Luo Q. (2019). Value of folate receptor-positive circulating tumour cells in the clinical management of indeterminate lung nodules: A non-invasive biomarker for predicting malignancy and tumour invasiveness. EBioMedicine, 41, 236-243.

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Measuring Senescence in Osteocytes

Cited 2 times

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Left hemi-jawbones were used to measure senescence-associated distension of satellites (SADS) in osteocytes located in cortical bone (n=10). For fluorescent in situ hybridization (FISH) staining, bone sections were processed as previously described [23 (link)]. The Cy3-labelled CENPB-specific (5’ ATTCGTTGGAAACGGGA 3’) peptide nucleic acid (PNA) FISH probe (Panagene Inc, Korea) was used, and slides were mounted with a DAPI-containing mounting media (ThermoFisher Scientific, Waltham, MA, USA). For SADS visualization, images were obtained using inverted laser scanning confocal microscope (LSM 780; Carl Zeiss Microscopy, Jena, Germany) with in-depth Z stacking. A senescent osteocyte was defined based on the number of SADS per cell, with a cut-off of ≥ 4 [23 (link), 24 (link)]. Nuclei of at least 50 osteocytes were analyzed for each sample.

Aquino-Martinez R., Rowsey J.L., Fraser D.G., Eckhardt B.A., Khosla S., Farr J.N, & Monroe D.G. (2020). LPS-Induced Premature Osteocyte Senescence: Implications in Inflammatory Alveolar Bone Loss and Periodontal Disease Pathogenesis. Bone, 132, 115220.

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Measuring Cell-Cell Fusion in BeWo Cells

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The cell–cell fusion of BeWo cells was detected and fusion index was calculated as described in Zhang et al. (2020 (link)) with some modifications. Briefly, forskolin was used to induce the syncytialization of BeWo cells. After being cultured for 24 h on poly-l-lysine-coated cover glass, BeWo cells with indicated treatments were treated with 30 μM forskolin (Cell Signaling Technology, USA) for 48 h. After the treatment, the culture medium was discarded and 4% paraformaldehyde was added to fix the cells by incubation for 20 min at room temperature. 0.5% Triton X-100 in PBS was used to permeabilize the cells for 3–5 min at room temperature. Blocking buffer was added to block the cells for 1 h at room temperature. Specific primary antibody anti-E-cadherin (Cell Signaling Technology, USA) was added in the blocking buffer and incubated with cells overnight at 4 °C. Primary antibody was washed out using PBS for 3 times, followed by incubation of Alexa fluor-conjugated secondary antibody for 2 h at room temperature. Cells were washed three times again by PBS and then treated with DAPI-containing mounting media (Thermo Fisher Scientific, USA). The cells were captured using fluorescence microscope (Zeiss, Germany).

Wu H.Y., liu K, & Zhang J.L. (2022). LINC00240/miR-155 axis regulates function of trophoblasts and M2 macrophage polarization via modulating oxidative stress-induced pyroptosis in preeclampsia. Molecular Medicine, 28, 119.

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Immunofluorescent Staining of SHSY5Y Cells

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SHSY5Y grown on 13mm coverslips cells were washed with PBS and fixed with 4% paraformaldegyde. The samples were permeabilized with PBS Triton X-100 (0.1%). Antibodies were prepared in 3% BSA. The calreticulin antibody was obtained from Abcam (AB2907) and the Goat Anti-chicken secondary was obtained from Invitrogen. The cover slips were mounted using DAPI-containing mounting media (Invitrogen) on pre-cleaned microscope slides and visualized on a Zeiss Axioscope confocal microscope.

Schiavon E., Smalley J.L., Newton S., Greig N.H, & Forsythe I.D. (2018). Neuroinflammation and ER-stress are key mechanisms of acute bilirubin toxicity and hearing loss in a mouse model. PLoS ONE, 13(8), e0201022.

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Fixation and Fluorescence Labeling of circMETTL3

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Four percent of PFA was added to Caco2 and HCT15 cells for fixation at room temperature for 10–12 min. PBS was used to wash off the PFA and the cells were then permeabilized with 1% Triton X-100 for 3–5 min at room temperature. Fluorescence-labeled specific probe for circMETTL3 was incubated with cells at 37 °C overnight in darkness. The probes were washed off by PBS and the cells were mounted on the slies with DAPI-containing mounting media (Invitrogen, USA).

Zhang F., Su T, & Xiao M. (2022). RUNX3-regulated circRNA METTL3 inhibits colorectal cancer proliferation and metastasis via miR-107/PER3 axis. Cell Death & Disease, 13(6), 550.

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Immunofluorescence Staining of HCT116 Cells

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HCT116 cells were cultured onto glass coverslips and grown for 24 h (75% confluence), washed twice with PBS and fixed for 20 min with 4% paraformaldehyde at room temperature. Then cells were washed with PBS and permeabilized with 0.1% saponin for 30 min in PBS, and subsequently blocked in 2% BSA blocking buffer for 1 h. Next, cells were stained with primary antibodies overnight at 4°C. Coverslips were rinsed five times with PBS and subsequently incubated with anti-Rabbit IgG Atto 488 (Sigma-Aldrich; #18772) and anti-Mouse IgG Atto 555 (Sigma-Aldrich; #43394) (1:200) for 1 h at room temperature followed by further five washes in PBS. Cells were then incubated with or without 100 nM Acti-stain™670 phalloidin (Cytoskeleton, Inc) for another 30 min. After washing with PBS five times, coverslips were mounted on glass slides by using DAPI-containing mounting media (Invitrogen) and analyzed by an LSM710 confocal microscope (Carl Zeiss AG).

Bacolla A., Sengupta S., Ye Z., Yang C., Mitra J., De-Paula R.B., Hegde M.L., Ahmed Z., Mort M., Cooper D.N., Mitra S, & Tainer J.A. (2020). Heritable pattern of oxidized DNA base repair coincides with pre-targeting of repair complexes to open chromatin. Nucleic Acids Research, 49(1), 221-243.

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Immunofluorescence Staining of G-Quadruplex DNA

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HeLa cells were purchased from ATCC and grown in DMEM medium (Corning) supplemented with 10% FBS (Sigma-Aldrich) and 1% (v/v) penicillin/streptomycin (Life Technologies) in a humidified incubator at 37°C with 5% CO₂. HAP1 cells were purchased from Horizon Discovery and cultured in Iscove’s modified Dulbecco’s medium (Corning) supplemented with 10% FBS (Sigma-Aldrich) and 1% (v/v) penicillin/streptomycin (Life Technologies) in a humidified incubator at 37°C with 5% CO₂. Cells were seeded onto glass coverslips and grown for 24 h, washed twice with PBS and fixed for 20 min with 4% paraformaldehyde at room temperature. After rinsing twice with PBS, cells were permeabilized for 30 min with 0.1% saponin in PBS, treated with RNase A (Roche) and subsequently blocked in 2% BSA blocking buffer for 1 h. Next, cells were incubated with the anti-G-quadruplex DNA 1H6 (Millipore) antibody (1:200) overnight at 4°C. Coverslips were rinsed four times with PBS and subsequently incubated with anti-Mouse IgG Atto 488 (Sigma-Aldrich) (1:200) for 1 h at room temperature followed by further four washes in PBS. Cells were then stained with 100 nM Acti-stain™ 670 phalloidin (Cytoskeleton, Inc) for 30 min followed by four washes in PBS. Coverslips were mounted on glass slides using DAPI-containing mounting media (Invitrogen) and analyzed using an LSM710 confocal microscope (Carl Zeiss AG).

Bacolla A., Ye Z., Ahmed Z, & Tainer J.A. (2019). Cancer mutational burden is shaped by G4 DNA, replication stress and mitochondrial dysfunction. Progress in biophysics and molecular biology, 147, 47-61.

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