6 diamidino 2 phenylindole dapi

Manufactured by Beyotime

Sourced in China, United States

6-diamidino-2-phenylindole (DAPI) is a fluorescent stain that binds strongly to adenine-thymine (A-T) rich regions in DNA. It is commonly used in fluorescence microscopy to visualize and identify cell nuclei.

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Lab products found in correlation

Triton x 100, by Merck Group (2 mentions) Utbi90 fluorescence microscope, by Olympus (2 mentions) Albumin from bovine serum bsa, by Thermo Fisher Scientific (1 mentions) Fitc conjugated goat anti mouse igg, by Beyotime (1 mentions) Dnmt1, by Affinity Biosciences (1 mentions) Ir 780 iodide, by Merck Group (1 mentions) Polyvinyl alcohol (pva), by Merck Group (1 mentions) Interleukin 4 (il 4), by BioLegend (1 mentions) Fluorescently labeled secondary antibody, by Abcam (1 mentions) Confocal laser scanning microscope, by Leica (1 mentions) Facscan flow cytometry, by BD (1 mentions) Cryostat, by Thermo Fisher Scientific (1 mentions) Fluorescence microscope, by Olympus (1 mentions) Ab26350, by Abcam (1 mentions) Paraformaldehyde (pfa), by MP Biomedicals (1 mentions) Alexa fluor 488 labeled goat anti mouse igg, by Beyotime (1 mentions) Cy3 labeled goat anti mouse igg, by Beyotime (1 mentions) Cy3 labeled goat anti rat igg, by Beyotime (1 mentions) Anti mannose receptor antibody, by Abcam (1 mentions) Bovine serum albumin (bsa), by Merck Group (1 mentions)

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14 protocols using 6 diamidino 2 phenylindole dapi

Immunohistochemical Analysis of Pancreatic Cell Proliferation

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Pancreases were freshly removed from the rats, 6-µm sections were cut
immediately, and fixed with acetone for hematoxylin-eosin (H&E) staining
using standard techniques. Some sections were subjected to immunohistochemical
staining (animals were previously intravenously injected with BrdU for 3 days).
After the process consisting of 10 min of fixation with acetone, 2 h of
permeabilization with 0.3% Triton X-100 (Sigma, St. Louis, MO, USA), and 1 h of
blocking with 3% albumin from bovine serum (BSA) (Gibco) at room temperature,
the sections were incubated with the primary antibody rabbit anti-rat BrdU
(1:100, Proteintech Group, Chicago, IL, USA) overnight at 4 °C, followed by a
further incubation with the secondary antibody goat anti-rabbit IgG-TRITC (1:50,
Proteintech Group) for 60 min at room temperature to detect the cycling cells in
the pancreas. 6-Diamidino-2-phenylindole (DAPI, Beyotime, Shanghai, China) was
used to stain the nuclei for 10 min. The pictures were captured using Immuno
Floure (Olympus, Tokyo, Japan).

Li G., Peng H., Qian S., Zou X., Du Y., Wang Z., Zou L., Feng Z., Zhang J., Zhu Y., Liang H, & Li B. (2020). Bone Marrow-Derived Mesenchymal Stem Cells Restored High-Fat-Fed Induced Hyperinsulinemia in Rats at Early Stage of Type 2 Diabetes Mellitus. Cell Transplantation, 29, 0963689720904628.

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Immunofluorescence Staining of Numb Protein

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The cells were fixed with 4% paraformaldehyde for 20 min, and then washed with PBS and permeabilization with 0.1% Triton-X 100. After blocking with 5% BSA for 30 min at room temperature, the cells were incubated with the anti-Numb antibody (1:150) at 4 °C overnight. Cells were rinsed with PBS for three times, and added with FITC-conjugated goat anti-mouse IgG (ZF-0311; Beijing, China, 1:200) for 1 h. 6-diamidino-2-phenylindole (DAPI) (C1005; Beyotime Institute of Biotechnology, Shanghai, China) was used to identify the nucleus. The images were captured by an Olympus UTBI90 Fluorescence microscope with the appropriate filters and identical acquisition parameters at ×100 magnification.

Zhang Z., Qu J., Zheng C., Zhang P., Zhou W., Cui W., Mo X., Li L., Xu L, & Gao J. (2018). Nrf2 antioxidant pathway suppresses Numb-mediated epithelial–mesenchymal transition during pulmonary fibrosis. Cell Death & Disease, 9(2), 83.

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Immunofluorescence Staining of Gastrocnemius Muscle

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The gastrocnemius muscles were fixed in 4% paraformaldehyde and embedded in paraffin. For immunofluorescence staining, the embedded samples were sectioned, and these sections (4 μm) were deparaffinized and rehydrated using dimethylbenzene, graded ethanol, and water baths. Antigens were retrieved using sodium citrate buffer for 10 min at 95 °C. The sections were washed three times with PBS. Afterward, the sections were permeabilized with 0.01% Triton X-100 for 15 min. After washing three times with PBS, the sections were incubated in PBS containing 5% goat serum for 2 h to avoid non-specific protein binding. The sections were incubated with primary CD31 (mouse monoclonal antibody, 1:100; Novus, Littleton, CO, USA) and/or DNMT1 (rabbit polyclonal antibody, 1:100; Affinity, USA) overnight at 4 °C and further stained with Alexa Fluor^® 488 anti-rabbit IgG (H+L) (1:100; CST, USA) and/or Alexa Fluor^® 594 anti-mouse IgG (H+L) (1:100; CST, Topsfield, MA, USA) for 2 h at room temperature. After washing three times with PBS, 6-diamidino-2-phenylindole (DAPI; Beyotime, Shanghai, China) was used to visualize nuclear localization. Finally, the sections were immediately examined using a confocal microscope (ZEISS, Jena, Germany). The data were analyzed using ZEN 2010.

Xue W., Zhang Q., Chen Y, & Zhu Y. (2022). Hydrogen Sulfide Improves Angiogenesis by Regulating the Transcription of pri-miR-126 in Diabetic Endothelial Cells. Cells, 11(17), 2651.

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Multifunctional IR780-Loaded Nanoparticles

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IR780 iodide, PLGA, and polyvinyl alcohol (PVA) were provided from Sigma Aldrich (St. Louis, MO, USA). Singlet Oxygen Sensor Green (SOSG) probe, IL-4, CD86 monoclonal antibody, CD80 monoclonal antibody, CD206 monoclonal antibody, and CD11c monoclonal antibody were purchased by Biolegend (San Diego, CA, USA). Cell-Counting Kit-8 (CCK-8), Calcein AM, 4, 6-diamidino-2-phenylindole (DAPI) and pyridine iodide (PI) were obtained from Beyotime Biotechnology (Shanghai, China). Staphylococcus aureus strains (MRSA, ATCC43300) and RAW 264.7 cell lines were obtained from Second Xiangya Hospital of Central South University (Changsha, China). All other reagents were of analytical grade and used without further purification.

Chen S., Wang J., Tang K., Liao H., Xu Y., Wang L, & Niu C. (2022). Multi-Modal Imaging Monitored M2 Macrophage Targeting Sono-Responsive Nanoparticles to Combat MRSA Deep Infections. International Journal of Nanomedicine, 17, 4525-4546.

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Immunostaining and Confocal Microscopy Protocol

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In total, 5 × 10⁴ cells were seeded into a confocal laser cuvette. After 24 h of cell climbing, 4% paraformaldehyde was fixed for 1 h, 0.25% Triton X-100 was ruptured for 1 h, and 5% bovine serum albumin was blocked for 1 h. Cells were immunostained with antibodies overnight at 4 °C, washed, and incubated with fluorescently labeled secondary antibodies (Abcam, MA, USA) at 37 °C for 1 h, and nuclei were stained with 6-diamidino-2-phenylindole (DAPI) (Beyotime, China). A confocal laser scanning microscope (Leica, Germany) was used for observation and imaging.

Gu J., Sun Y., Song J., Zhao R., Di X., Zhang Y., Ge X., Zhang S., Gu Y, & Sun X. (2022). Irradiation induces DJ-1 secretion from esophageal squamous cell carcinoma cells to accelerate metastasis of bystander cells via a TGF-β1 positive feedback loop. Journal of Experimental & Clinical Cancer Research : CR, 41, 259.

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Apoptosis Quantification by Annexin V/PI Staining

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Apoptosis of cells incubated in 1% FBS was determined by staining with fluorescein
isothiocyanate (FITC)-labeled annexin V and propidium iodide (PI; PharMingen, USA),
according to the manufacturer's instructions. The cells were stained simultaneously
with FITC-labeled annexin V and PI and scored as follows: 1) annexin V-/PI- (viable
cells); 2) annexin V+/PI- (cells in the initial stages of apoptosis); 3) annexin
V+/PI+ (cells in the advanced stages of apoptosis), and 4) annexin V-/PI+ (necrotic
cells). To quantify apoptosis, the cells were washed with cold PBS and then suspended
in binding buffer. The cells were stained with 5 µL annexin V-FITC and 10 µL PI and
then analyzed using FACScan flow cytometry (FCM; Becton Dickinson, USA) at 48 h.
4′,6-Diamidino-2-phenylindole (DAPI, Beyotime) was added to the culture medium to
determine morphological changes during apoptosis; and the fragmentation of the
nucleus and chromatin condensation were examined by fluorescence microscopy.
The three groups (CEP-caspase-3, CEP-NC, and CEP-CTR) at a density of
1×10⁵ cells/well were incubated in 1% FBS for 24 and 48 h, and
harvested after trypsinization. Apoptosis of cells transfected with EGFP and control
group cells was determined by staining with annexin V phycoerythrin (PE) (Beyotime),
and analyzed using FCM.

Ding L., Wu J.P., Xu G., Zhu B., Zeng Q.M., Li D.F, & Lu W. (2014). Lentiviral-mediated RNAi targeting caspase-3 inhibits apoptosis induced by serum deprivation in rat endplate chondrocytes in vitro. Brazilian Journal of Medical and Biological Research, 47(6), 445-451.

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Immunofluorescence Assay for Nrf2 Expression

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The cells were fixed with 4% paraformaldehyde for 20 minutes and then washed with PBS and permeabilization with 0.1% Triton ×100. After blocking with 5% BSA for 30 minutes at room temperature, the cells were incubated with the anti‐Nrf2 antibody (1:200) at 4°C overnight. Cells were rinsed with PBS for 3 times and added with FITC‐conjugated goat anti‐rabbit IgG (ZF‐0311; Beijing, China, 1:200) for 1 hour. 6‐diamidino‐2‐phenylindole (DAPI) (C1005; Beyotime Institute of Biotechnology, Shanghai, China) was employed to identify the nucleus. The images were captured by an Olympus UTBI90 Fluorescence microscope with the appropriate filters and identical acquisition parameters at ×200 magnification.

Cui W., Zhang Z., Zhang P., Qu J., Zheng C., Mo X., Zhou W., Xu L., Yao H, & Gao J. (2018). Nrf2 attenuates inflammatory response in COPD/emphysema: Crosstalk with Wnt3a/β‐catenin and AMPK pathways. Journal of Cellular and Molecular Medicine, 22(7), 3514-3525.

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Cerebral Cortex and Retinal Tissue Analysis

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The cerebral cortex and eyeballs were dissected after MCAO treatment. The eyeballs were fixed in Fekete's solution for 1 h. The cornea, lens, and vitreous humor were removed. Then, cortex and retinas were fixed in 4% PFA overnight at 4 °C. Next, they were dehydrated in a gradient sucrose solution (15% and 30%) at 4 °C, followed by embedding in OCT compound. Serial sections of cerebral cortex (20 μm thick) and eye tissue (5 μm thick) were cut using a cryostat (Thermo Scientific, USA) and placed on adhesion microscope slides (Citotest). After washing twice with PBS, the frozen sections were permeabilized and blocked with 5% bovine serum albumin (BSA) for 30 min. Then, they were incubated with the primary antibody overnight at 4 °C and washed with PBST three times. The slides were incubated with the secondary antibody for 2 h and 4, 6-diamidino-2-phenylindole (DAPI, 1:1500, Beyotime, C1002) for 10 min in dark. The sections were observed under a fluorescent microscope (Olympus IX73).

Jiang Q., Su D.Y., Wang Z.Z., Liu C., Sun Y.N., Cheng H., Li X.M, & Yan B. (2021). Retina as a window to cerebral dysfunction following studies with circRNA signature during neurodegeneration. Theranostics, 11(4), 1814-1827.

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Quantifying DNA Damage using Immunofluorescence

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The fluorescence of cells was observed in a dark room using a fluorescence microscope. The treated cells were fixed in 4% paraformaldehyde for at least 20 min, washed three times using PBS, permeabilized using 0.2% Triton for 15 min, followed by washing three times using PBS, then sealed at 4°C with 5% goat serum (diluted by PBS) and incubated overnight with an anti-γ-H2AX antibody (ab26350; Abcam, USA). Under dark conditions, the cells were incubated with secondary antibodies for 2 h (diluted 1:200). The cells were covered with 6-diamidino-2-phenylindole (DAPI) (Beyotime) for 10 min for nuclear staining, and then immediately observed under an Olympus fluorescence microscope (Olympus America Inc., USA).

Wang K., Zhu Q.Z., Ma X.T, & Cheng C. (2021). SUV39H2/KMT1B Inhibits the cardiomyocyte senescence phenotype by down-regulating BTG2/PC3. Aging (Albany NY), 13(18), 22444-22458.

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Macrophage Polarization via Immunofluorescence

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Macrophage polarization (M0, M1, and M2) was identified by fluorescence staining and imaging. The three groups of cells were fixed for 30 min at room temperature in 4% paraformaldehyde (MP Biomedicals, Santa Ana, CA, United States). 0.1% Triton X-100 (Sigma-Aldrich, St. Louis, MO, United States) in PBS containing 0.1% Tween-20 was used to permeabilize the cells, after washing them with PBS thrice (5 min each). The cells were blocked with 3% BSA (Sigma-Aldrich) for 30 min at room temperature. After that, samples were allowed to incubate overnight at 4°C with primary antibodies, which included rabbit anti-iNOS IgG (Abcam, United States) and mouse anti-Arginase (Abcam, United States), Anti-CD86 Antibody (Abcam, United States), and Anti-Mannose Receptor Antibody (Abcam, United States) (Abcam, United States). Secondary antibodies included Cy3-labeled Goat Anti-Rat IgG (Beyotime, Shanghai, China), Cy3-labeled Goat Anti-Mouse IgG (Beyotime, Shanghai, China), and Alexa Fluor 488-labeled Goat Anti-Mouse IgG (Beyotime, Shanghai, China), and cells were counterstained with 6-diamidino-2-phenylindole (DAPI) (Beyotime, Shanghai, China) solution at 1 μg/ml. Images were obtained using a fluorescence microscope was used to capture the images (BX51; Olympus, Tokyo, and Japan).

Liu K., Luo X., Lv Z.Y., Zhang Y.J., Meng Z., Li J., Meng C.X., Qiang H.F., Hou C.Y., Hou L., Liu F.Z, & Zhang B. (2022). Macrophage-Derived Exosomes Promote Bone Mesenchymal Stem Cells Towards Osteoblastic Fate Through microRNA-21a-5p. Frontiers in Bioengineering and Biotechnology, 9, 801432.

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