4 6 diamidino 2 phenylindole (dapi)

Manufactured by Antgene

Sourced in China

DAPI is a fluorescent stain that binds to DNA. It is commonly used in microscopy and flow cytometry applications to visualize and quantify nucleic acids.

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Lab products found in correlation

Confocal microscope, by Olympus (3 mentions) Bx53 microscope, by Olympus (3 mentions) Rabbit anti sirt6 polyclonal antibody, by Thermo Fisher Scientific (2 mentions) Fluorescence microscope, by Olympus (2 mentions) Ant024s, by Antgene (2 mentions) Alexa fluor labelled secondary antibody, by Antgene (2 mentions) Bx51 microscope, by Olympus (2 mentions) Confocal laser scanning microscope, by Olympus (2 mentions) Alexa fluor 488 donkey anti mouse igg h l, by Jackson ImmunoResearch (1 mentions) Anti yap, by Cell Signaling Technology (1 mentions) Bovine serum albumin (bsa), by Antgene (1 mentions) Ab178518, by Abcam (1 mentions) Sc 515842, by Santa Cruz Biotechnology (1 mentions) Image pro plus v6, by Media Cybernetics (1 mentions) Fluorescent microscope, by Olympus (1 mentions) Eight well chamber slide, by Ibidi (1 mentions) Donkey serum, by Antgene (1 mentions) Fluorescent secondary antibody, by Proteintech (1 mentions) Neuronal nuclei (neun), by Proteintech (1 mentions) Caspase 1, by Novus Biologicals (1 mentions)

Spelling variants of this product

4′,6′-diamidino-2-phenylindole (DAPI, (3 citations)

20 protocols using 4 6 diamidino 2 phenylindole (dapi)

Immunostaining of Sirt6 and WT1 in Kidney Sections

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The paraffin-embedded kidney sections were first dewaxed and subjected to antigen retrieval. Then, they were blocked with 5% bovine serum albumin for 30 min at room temperature. The sections were next incubated with a mixture of rabbit anti-Sirt6 polyclonal antibody (1:100, Thermo Fisher Scientific, USA) and mouse anti-WT1 monoclonal antibody (1:100, Novus, USA) overnight at 4°C.The sections were stained with a mixture of Alexa Fluor 488, Donkey anti-mouse IgG (HL) (1:200, Jackson Immuno Research Laboratories, USA) and Alexa Fluor 594, Donkey anti-Rabbit IgG (HL) (1:200, Jackson Immuno Research Laboratories, USA) as the secondary antibodies at 37°C for 60 min in the dark. The nuclei were counterstained with DAPI (Antgene, China) for 5 min.
The cell-climbing films (cells growing on glass slides) were fixed with 4% paraformaldehyde for 30 min at 4°C and stained with rabbit anti-Sirt6 polyclonal antibody (1:100, Thermo Fisher Scientific, USA) overnight at 4°C. The films were then incubated with Alexa Fluor 594, Donkey anti Rabbit IgG (HL) (1:200, Jackson Immuno Research Laboratories, USA) at 37°C for 60 min. The nuclei were counterstained with DAPI (Antgene, China) for 5 min. A confocal microscope (Olympus, Japan) was used to record all of the microscopic images.

Fan Y., Yang Q., Yang Y., Gao Z., Ma Y., Zhang L., Liang W, & Ding G. (2019). Sirt6 Suppresses High Glucose-Induced Mitochondrial Dysfunction and Apoptosis in Podocytes through AMPK Activation. International Journal of Biological Sciences, 15(3), 701-713.

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Immunofluorescence Staining of Podocytes

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The paraffin sections were first dewaxed and then subjected to antigen retrieval. The slides were blocked for 30 min with 5% bovine serum albumin (BSA) and then incubated overnight at 4 °C with anti-synaptopodin (1:100, Santa, USA) and anti-ALCAT1 (1:100, Novus, USA) antibodies. On the next day, a mixture of fluorescent secondary antibodies was applied to the slides and incubated at room temperature in darkness for 1 h. DAPI (Antgene, China) was used to stain the nuclei for 5 min. Cultured podocytes were fixed on cover slides in 4% paraformaldehyde at 4 °C for half an hour and then incubated with ALCAT1 antibody (1:100, Novus, USA) overnight at 4 °C. Finally, the nuclei were stained with DAPI (Antgene, China). Images were captured using a confocal microscope (Olympus, Japan).

Hao Y., Fan Y., Feng J., Zhu Z., Luo Z., Hu H., Li W., Yang H, & Ding G. (2024). ALCAT1-mediated abnormal cardiolipin remodelling promotes mitochondrial injury in podocytes in diabetic kidney disease. Cell Communication and Signaling : CCS, 22, 26.

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Analyzing Ki67 Expression in PER2 Knockout CCD 841 CoN Cells

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To observe the effect of PER2 knockout on the expression level of Ki67 in the CCD 841 CoN cells, we cultured the same number of cells transfected with siRNA against PER2 (siPer2) and control siRNA (siCon) on 12-well slides overnight. On the second day, 4% paraformaldehyde was used to fix the cell slides for 15 min. They were washed three times with PBS, incubated with 1% donkey serum for 1 h, and then incubated with polyclonal rabbit anti-Ki67 antibody (1:200; cat. GB111499; Servicebio) at 4°C overnight. The next day, the sections were incubated with Alexa Fluor 594 anti-rabbit secondary antibody (1:200; cat. ANT029S; AntGene, Wuhan, Hubei, China) for 1 h at room temperature and then counterstained with DAPI (cat. ANT165; AntGene). The expression of Ki67 was examined using a laser scanning confocal microscope (Si-A1; Nikon, Tokyo, Japan).

Chen Y.D., Zhao R.F., Zheng G., Ling F.M., Li J.R., Xu M.Y., Guo D., Zhang Q.L., Li S, & Zhu L.R. (2022). The association between disruption of the circadian rhythm and aggravation of colitis in mice. Gastroenterology Report, 10, goac028.

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Immunohistochemical analysis of JMJD3 and YAP in colon tissue

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The Paraffin-embedded colonic tissue sections were deparaffinized, blocked with 5% bovine serum albumin (BSA) for 1 h, and then incubated with primary antibodies at 4 °C overnight (anti-JMJD3, 1:50, Abclonal, A17382; anti-YAP, 1:50, Cell Signaling Technology, 14,074). After washing with PBS, the sections were incubated with secondary antibodies (Thermo Fisher Scientific) at 37 °C for 2 h in the dark. Nuclei were counterstained with DAPI (Antgene). All images were obtained using a confocal microscope (Olympus, Tokyo, Japan).

Zhu H., Lu J., Fu M., Chen P., Yu Y., Chen M., Zhao Q., Wu M, & Ye M. (2024). YAP represses intestinal inflammation through epigenetic silencing of JMJD3. Clinical Epigenetics, 16, 14.

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Immunofluorescence Staining of Kidney and Cell Cultures

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Following fixation with 4% paraformaldehyde for 30 min and permeabilization in 0.2% Triton X-100 (PBS) at 4°C, the frozen kidney sections were blocked with 5% bovine serum albumin (Antgene) at room temperature for 1 h. The sections were then incubated with a mixture of primary antibodies against sesn2 rabbit monoclonal antibody (1:100; ab178518, Abcam) and mouse anti-snaptopodin (1:200; sc-515842, Santa Cruz Biotechnology, Inc.) overnight at 4°C. The sections were subsequently stained with a mixture of Alexa Fluor 488, donkey anti-rabbit IgG (HL) (1:200; ANT024s, Antgene) and Alexa Fluor 594, donkey anti-Mouse IgG (HL) (1:200; ANT029s, Antgene) secondary antibodies at 37°C for 60 min in the dark. The nuclei were counterstained with DAPI (Antgene) at 37°C for 5 min.
The cell climbing films were fixed in 4% paraformaldehyde for 20 min at 4°C, and blocked with 5% bovine serum albumin (Antgene) at room temperature for 1 h. The films were then stained with the following antibodies at 4°C overnight: Sesn2 rabbit monoclonal antibody (1:100; ab178518, Abcam). The films were then incubated with Alex Fluor 488, donkey anti-rabbit IgG (HL) (1:200; ANT024s, Antgene) at 37°C for 60 min in the dark. The nuclei were counterstained with DAPI 37°C for 5 min. All microscopic images were visualized with a fluorescence microscope (Olympus Corp.).

Lin Q., Ma Y., Chen Z., Hu J., Chen C., Fan Y., Liang W, & Ding G. (2020). Sestrin-2 regulates podocyte mitochondrial dysfunction and apoptosis under high-glucose conditions via AMPK. International Journal of Molecular Medicine, 45(5), 1361-1372.

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Immunostaining of PDGFR and TGFβ1-R in gb-MSCs

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gb-MSCs (5,000 cells/well) were cultured in an eight-well chamber slide (Ibidi GmbH) at 37°C. Once cells reached 70% confluence, they were washed with PBS three times, fixed with 4% paraformaldehyde for 15 min at room temperature, permeabilized with 0.5% Triton-X-100 for 20 min at room temperature and blocked with a solution containing donkey serum (Antgene) for 1 h at room temperature. Subsequently, cells were incubated with primary antibodies against PDGFR and TGFβ1-R (R&D Systems, Inc.) overnight at 4°C. After being washed three times with PBS, cells were incubated with secondary antibodies labeled with rhodamine (Antgene) for 1 h at room temperature. In each experiment, non-specific staining by secondary antibodies was excluded by incubating a well without primary antibodies. Cells were also stained with DAPI for 5 min at room temperature (Antgene). Images were acquired using a fluorescent microscope (Olympus Corporation) and data were analyzed using Image-Pro Plus v6.0 (Media Cybernetics, Inc.).

Zhang Q., Xiang W., Xue B.Z., Yi D.Y., Zhao H.Y, & Fu P. (2021). Growth factors contribute to the mediation of angiogenic capacity of glioma-associated mesenchymal stem cells. Oncology Letters, 21(3), 215.

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Immunofluorescence Visualization of Cells

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The sterilized slide was placed in 6-well plates with about 3 × 10⁴ cells/well. Following 12-h treatment with DDP, cells were fixed for 30 min in 4% paraformaldehyde, then permeated for 10 min in 0.5% Triton X-100, and sealed for 30 min using 1% albumin bovine V (ServiceBio, Changchun, Jilin, China). The cells were subjected to overnight incubation with primary antibody at 4°C and 1-h incubation with secondary antibody (Antgene, Wuhan, Hubei, China) in the dark. Subsequently, the nuclei were dyed with DAPI (Antgene) under dark conditions. Images were acquired using an automatic microscope (BX63, Olympus).

Zheng J., Li X., Cai C., Hong C, & Zhang B. (2021). MicroRNA-32 and MicroRNA-548a Promote the Drug Sensitivity of Non-Small Cell Lung Cancer Cells to Cisplatin by Targeting ROBO1 and Inhibiting the Activation of Wnt/β-Catenin Axis. Cancer Management and Research, 13, 3005-3016.

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Immunofluorescent Staining of Apoptosis Markers

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The cells were fixed with 4% paraformaldehyde for 30 min, ruptured with 0.5% Triton X-100 for 10 min, and blocked with 1% BSA for 30 min. Then, the cells were incubated with the diluted primary antibodies (including anti-BCL2 and anti-cleaved caspase 3) overnight at 4 °C. The next day, the cells were incubated with Alexa Fluor-labelled secondary antibodies (Antgene, China) for 1 h under dark conditions. The nuclei were stained with DAPI (Antgene, China) in the dark for 5 min. An Olympus BX51 microscope (Olympus, Japan) was used to capture the images.

Cai J., Gao L., Wang Y., Li Y., Ye Z., Tong S., Yan T., sun Q., Xu Y., Jiang H., Zhang S., Zhao L., Yang J, & Chen Q. (2022). TMBIM1 promotes proliferation and attenuates apoptosis in glioblastoma cells by targeting the p38 MAPK signalling pathway. Translational Oncology, 19, 101391.

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Immunohistochemical Analysis of Neural Apoptosis

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Brain paraffin sections (4 μm) were hydrated and Tris/EDTA buffer performed heat-mediated antigen retrieval for 20 min. The sections blocked with 5% BSA for 1 h were incubated with Neun (Proteintech) along with primary antibodies Caspase-1 (Novus) overnight at 4°C and subsequently incubated in fluorescent secondary antibodies (Proteintech) for 1 h at 24°C. DAPI (Antgene) was used for nuclei staining. Images were taken with an Olympus BX53 microscope (Olympus). Positive cells were counted using ImageJ software (n = 6/group).

Liu J., Zheng J., Xu Y., Cao W., Wang J., Wang B., Zhao L., Zhang X, & Liao W. (2021). Enriched Environment Attenuates Pyroptosis to Improve Functional Recovery After Cerebral Ischemia/Reperfusion Injury. Frontiers in Aging Neuroscience, 13, 717644.

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Kidney Immunofluorescence Imaging Protocol

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The frozen kidney sections were blocked and incubated with a mixture of guinea pig anti-adipocyte differentiation-related protein (Adrp) antibody (1:100, Progen Biotechnik, Germany) and mouse anti-WT1 antibody (1:100, Novus, USA) or a mixture of rabbit anti-Sirt6 antibody (1:100, Thermo Fisher Scientific, USA) and mouse anti-WT1 antibody (1:100, Novus, USA)/mouse anti-Synaptopodin antibody (1:100, Progen Biotechnik, Germany) overnight at 4 °C, followed by incubation with fluorescent secondary antibodies (1:200, Thermo Fisher Scientific, USA) at 37 °C for 90 min in the dark. Nuclei were counterstained with DAPI (Antgene, China) for 5 min. All microscopic images were recorded with fluorescence microscope (Olympus, Japan).

Yang Q., Hu J., Yang Y., Chen Z., Feng J., Zhu Z., Wang H., Yang D., Liang W, & Ding G. (2020). Sirt6 deficiency aggravates angiotensin II-induced cholesterol accumulation and injury in podocytes. Theranostics, 10(16), 7465-7479.

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