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PUREBLU™ DAPI

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PUREBLU™ DAPI thumbnail image 1 PUREBLU™ DAPI thumbnail image 2 PUREBLU™ DAPI thumbnail image 3 PUREBLU™ DAPI thumbnail image 4 PUREBLU™ DAPI thumbnail image 5 PUREBLU™ DAPI thumbnail image 6 PUREBLU™ DAPI thumbnail image 7 PUREBLU™ DAPI thumbnail image 8 PUREBLU™ DAPI thumbnail image 9 PUREBLU™ DAPI thumbnail image 10
PUREBLU™ DAPI gallery image 1
HeLa cells supplemented with BrdU in vitro for 3 hours then stained with Mouse anti-BrdU antibody, clone Bu20a (MCA2483). Goat anti mouse IgG (H/L) DyLight® 549 conjugated antibody (red) (STAR117D549GA) was used as the secondary antibody. Cytoplasm was stained with Rabbit anti-GAPDH antibody (AHP1628). As the secondary antibody, Sheep anti Rabbit IgG DyLight 488 conjugated antibody (green) was used. PureBlu DAPI (1351303) was used as nuclear counterstain (blue).
PUREBLU™ DAPI gallery image 2
HeLa cells, formaldehyde fixed and permeabilized, stained with Rat anti alpha tubulin antibody, clone YOL1/34 (red) (MCA78G), Mouse anti Human CD49a antibody, clone TS2/7, green (MCA1133) and PureBlu DAPI, blue (1351303) as a nuclear counterstain.
PUREBLU™ DAPI gallery image 3
Paraffin section of human pancreas stained with Guinea Pig anti insulin antibody, clone D3e7 (red) (5330-0104G),and Mouse anti chromogranin A antibody, clone LK2H10 (MCA5696), green and PureBlu DAPI, blue (1351303) as a nuclear counterstain.
PUREBLU™ DAPI gallery image 4
Paraffin section of human colon adenocarcinoma (HIER citrate pH6 antigen retrieval) blocked with 10% FCS and stained with Mouse anti Human cytokeratin 18 antibody (VMA00314), green and detected with Goat anti Mouse IgG:A488 (STAR117D488GA). The tissue was also counterstained with PureBlu DAPI, blue (1351303).
PUREBLU™ DAPI gallery image 5
HeLa cells treated with 10 μg BrdU in vitrofor 1 hour (B) or left untreated (A). Cells were stained with Mouse anti BrdU antibody, clone Bu20a (MCA2483) at a dilution of 1/25. As a secondary antibody, goat anti mouse IgG (H/L) DyLight 549 conjugated antibody, red (STAR117D549GA) was used at a 1/50 dilution. Cytoplasm was stained with rabbit anti-GAPDH antibody (AHP1628) at a dilution of 1/100. As a secondary antibody, sheep anti rabbit IgG DyLight 488 conjugated antibody, green was used at a 1/50 dilution. PureBlu DAPI, blue (1351303) was used as nuclear counterstain.
PUREBLU™ DAPI gallery image 6
Published customer image:
PureBlu DAPI fluorescent DNA binding reagent (1351303) used to visualize nuclei of human circulating tumor cells by immunofluorescence in combination with cell surface markers.
Image caption:
Confocal microscopy representative images of isolated CTCs stained for epithelial and hematopoietic markers. Representative image by confocal microscopy of circulating cancer cells stained with anti-human EPCAM, anti-human pan-CK or anti-human CD45 Abs followed by goat anti-mouse secondary antibody Alexa 594-conjugated. DAPI was used to counteract nuclei. Magnification 100×. CK = cytokeratin, an epithelial cytoplasmic marker; DAPI = nuclear marker; EpCAM = epithelial membrane marker; CD45 = leukocyte common antigen.

From: Morelli MB, Amantini C, Rossi de Vermandois JA, Gubbiotti M, Giannantoni A, Mearini E, Maggi F, Nabissi M, Marinelli O, Santoni M, Cimadamore A, Montironi R, Santoni G.
Correlation between High PD-L1 and EMT/Invasive Genes Expression and Reduced Recurrence-Free Survival in Blood-Circulating Tumor Cells from Patients with Non-Muscle-Invasive Bladder Cancer.
Cancers (Basel). 2021 Nov 28;13(23):5989.
doi: 10.3390/cancers13235989.
This image is from an open access article distributed under terms of a Creative Commons Attribution License.
PUREBLU™ DAPI gallery image 7
Published customer image:
PureBlu DAPI fluorescent DNA stain (1351303) used to stain nuclei of RAW 264.7 cells by immunofluorescence.
Image caption:
Effect of astaxanthin on F-actin ring size. (A) Cells were treated similarly to the TRAP activity experiment, which was followed by staining with phalloidin to stain cytoplasm as well as DAPI to stain nuclei. F-actin rings are indicated by white arrows in the figures. (B) F-actin ring size per osteoclast cells. All data are shown as means ±SEM, ** p <0.01, Tukey–Kramer test. The bar in the figure represents 50μm.

From: Mamun-Or-Rashid, A.N.M.; Lucy, T.T.; Yagi, M.; Yonei, Y.
Inhibitory Effects of Astaxanthin on CML-HSA-Induced Inflammatory and RANKL-Induced Osteoclastogenic Gene Expression in RAW 264.7 Cells.
Biomedicines 2022, 10, 54.
doi: 10.3390/biomedicines10010054
This image is from an open access article distributed under terms of a Creative Commons Attribution License.
PUREBLU™ DAPI gallery image 8
Cultured Jurkat cells harvested, washed in phosphate buffered saline and fixed in cold 70% ethanol at 4°C for 2 hours. Cells were then washed in Staining Buffer (BUF073) and stained for 30 min with a solution of PureBlu DAPI (1351303) containing 0.1% Triton X-100. Cells were first gated to discriminate single cells and then plotted into histograms discriminating cells in G1, S and G2/M phases.
PUREBLU™ DAPI gallery image 9
Flow cytometry analysis of Jurkat cells heat shocked at 65°C for 5 min and stained with PureBlu DAPI (1351303). Dead cells show positive staining.
PUREBLU™ DAPI gallery image 10
Published customer image:
PureBlu DAPI Nuclear stain used to highlight nuclei in RAW264.7 cells in vitro by immunofluorescence.
Image caption:
Effect of passage number on F-actin ring formation. (A) Cells were seeded and treated same as the TRAP staining experiment followed by staining with phalloidin to stain the cytoplasm (green), an antibody against RAGE (red), and DAPI (blue) to stain the nucleus. The bar in the figure represents 100 μm. White arrows indicate osteoclast cells. (B) F-actin ring number in two different passages. ** p <0.01, Tukey–Kramer test.

From: Lucy TT, Mamun-Or-Rashid ANM, Yagi M, Yonei Y.
Serial Passaging of RAW 264.7 Cells Modulates Intracellular AGE Formation and Downregulates RANKL-Induced In Vitro Osteoclastogenesis.
Int J Mol Sci. 2022 Feb 21;23(4):2371.
doi: 10.3390/ijms23042371.
This image is from an open access article distributed under terms of a Creative Commons Attribution License.

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PUREBLU™ DAPI

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Accessory Reagent
Specificity
PUREBLU™ DAPI

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Product Code Applications Pack Size List Price Your Price Qty
1351303
Datasheet Datasheet Datasheet
SDS Safety Datasheet SDS
F IF 250 µg
List Price Your Price

DAPI is a cell permeable fluorescent compound (MW 350.3) that is able to stain the DNA of eukaryotic and prokaryotic cells by binding with high affinity to the minor groove of AT-rich DNA sequences. When DAPI is bound to DNA and excited by an ultraviolet light source, blue fluorescent emission can be detected with maximum emission at 461 nm. PureBlu DAPI has a characteristic Stokes shift of approximately 100 nm, which makes this dye an optimal choice when good spectral separation is desired. PureBlu DAPI is compatible with fixed and unfixed cells.

Reconstitution
Reconstitute one vial of lyophilized PureBlu DAPI Dye with 500 μl of de-ionized water, then vortex briefly to make a 100x stock solution.
Reagents in the Kit
5 vials, 50 μg each, DAPI nuclear staining dye powder
Regulatory
For research purposes only
Guarantee
Guaranteed until date of expiry. Please see product label.
Acknowledgements
Triton is a trademark of Dow Chemical Company

Prior to reconstitution store at - 20°C
After reconstitution store at +4°C or at -20°C if preferred
This product is photosensitive and should be protected from light. PureBlu DAPI Dye is stable for 12 months from date of reconstitution if stored at - 20°C or 6 months at 2-8°C.

This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit the antibody protocols page.
Application Name Verified Min Dilution Max Dilution
Flow Cytometry 1/100
Immunofluorescence 1/100
Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
Instructions For Use
Note: The optimal concentration for different cell types should be determined empirically.

Staining Procedure - Flow Cytometry Cell Viability

1. Grow cells of interest under conditions specific for the cell type. Harvest cells and obtain a single cell suspension.

2. Check cell numbers and viability using trypan blue.

3. Resuspend cells at 2 x 106 cells/ml in 0.5ml Staining Buffer (BUF073) or 1x PBS.

4. Add 5μl of 100x PureBlu DAPI Dye stock solution to 0.5ml cells.

5. Stain at room temperature for 15 min in the dark.

6. Optional: Rinse cells with Staining Buffer (BUF073) or 1x PBS.

7. Proceed with analysis by flow cytometry. When bound to double-stranded DNA (dsDNA) PureBlu DAPI Dye can be excited with either a 355 nm or a 405 nm laser, with optimum emission at 461nm.


Staining Procedure - Flow Cytometry Cell DNA Content

1. Grow cells of interest under conditions specific for the cell type. Harvest cells and obtain a single cell suspension.

2. Check cell numbers and viability using trypan blue.

3. Wash cells in Staining Buffer (BUF073) or 1x PBS.

4. Fix in ice cold 70% ethanol for 2 hr at 4°C.

5. Wash cells in Staining Buffer (BUF073) or 1x PBS.

6. Stain cells with 5μl of PureBlu DAPI Dye stock solution to 0.5 ml cells at a concentration of 2 x 106 cells/ml in cell staining buffer containing 0.1% Triton X-100.

Note: Optimal concentrations of cells and PureBlu DAPI may vary depending on cell type and should be determined through careful titration prior to final experimental analysis.

7. Stain at room temperature for 30 min in the dark.

8. Optional: Rinse cells with Staining Buffer (BUF073) or 1x PBS.

9. Proceed with analysis by flow cytometry. When bound to double-stranded DNA (dsDNA) PureBlu DAPI Dye can be excited with either a 355 nm or a 405 nm laser, with optimum emission at 461nm.


Staining Procedure - Cell Nuclear Visualization in Microscopy and Cell Imaging

Preparation of the 1x staining solution: Dilute the 100x stock solution 1/100 with PBS for a final concentration of 1 μg/ml.

1. Grow cells of interest under conditions specific for the cell type.

2. Rinse cells with 1x PBS.

3. Optional: Rinse cells with 1x PBS and permeabilize them with 1x PBST (0.1% Triton X-100 in 1x PBS) at room temperature for 5 min.

5. Rinse cells with 1x PBS.

6. Stain with 1x staining solution (diluted with PBS) at room temperature for 15 min.

7. Rinse cells with 1x PBS.

8. Optional: Remove PBS and mount cells in antifade mounting media.

9. Image cells.

Description Product Code Applications Pack Size List Price Your Price Quantity
Staining Buffer BUF073 F 500 ml
List Price Your Price
Description Staining Buffer

References for PUREBLU™ DAPI

  1. She, D.T. et al. (2018) SIRT2 Inhibition Confers Neuroprotection by Downregulation of FOXO3a and MAPK Signaling Pathways in Ischemic Stroke.
    Mol Neurobiol. 55 (12): 9188-203.
  2. Carter-Timofte, M.E. et al. (2021) Antiviral Potential of the Antimicrobial Drug Atovaquone against SARS-CoV-2 and Emerging Variants of Concern.
    ACS Infect Dis. acsinfecdis.1c00278.
  3. Earnest, K.G. et al. (2021) Development and characterization of a DNA aptamer for MLL-AF9 expressing acute myeloid leukemia cells using whole cell-SELEX.
    Sci Rep. 11 (1): 19174.
  4. Paterno, R. et al. (2021) Hippocampal gamma and sharp-wave ripple oscillations are altered in a Cntnap2 mouse model of autism spectrum disorder.
    Cell Rep. 37 (6): 109970.
  5. Roux-Biejat, P. et al. (2021) Acid Sphingomyelinase Controls Early Phases of Skeletal Muscle Regeneration by Shaping the Macrophage Phenotype.
    Cells. 10 (11): 3028.
  6. Nogueira-Rodrigues, J. et al. (2021) Rewired glycosylation activity promotes scarless regeneration and functional recovery in spiny mice after complete spinal cord transection.
    Dev Cell. Dec 27 [Epub ahead of print].
  7. Morelli, M.B. et al. (2021) Correlation between High PD-L1 and EMT/Invasive Genes Expression and Reduced Recurrence-Free Survival in Blood-Circulating Tumor Cells from Patients with Non-Muscle-Invasive Bladder Cancer.
    Cancers (Basel). 13 (23)Nov 28 [Epub ahead of print].
  8. Mamun-Or-Rashid, A.N.M. et al. (2021) Inhibitory Effects of Astaxanthin on CML-HSA-Induced Inflammatory and RANKL-Induced Osteoclastogenic Gene Expression in RAW 264.7 Cells
    Biomedicines. 10 (1): 54.
    9. View The Latest Product References
  9. Saito, N. et al. (2022) Impaired dental implant osseointegration in rat with streptozotocin-induced diabetes.
    J Periodontal Res. Jan 17 [Epub ahead of print].
  10. Lucy, T.T. et al. (2022) Serial Passaging of RAW 264.7 Cells Modulates Intracellular AGE Formation and Downregulates RANKL-Induced In Vitro. Osteoclastogenesis.
    Int J Mol Sci. 23 (4)Feb 21 [Epub ahead of print].
  11. Foo, S.L. et al. (2022) Breast cancer metastasis to brain results in recruitment and activation of microglia through annexin-A1/formyl peptide receptor signaling.
    Breast Cancer Res. 24 (1): 25.
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