Thermo Scientific™ DAPI (4',6-diamidino-2-phenylindole, dihydrochloride)

Catalog No.	Quantity
PI62247	10 mg

Catalog No. PI62247 Supplier Thermo Scientific™ Supplier No. 62247

Description

Thermo Scientific Pierce DAPI Nuclear Counterstains are high-purity forms of diamidino-2-phenylindole dye for fixed-cell, fluorescent staining of DNA content and nuclei for cellular imaging techniques.

Thermo Scientific™ Pierce DAPI Nuclear Counterstains are high-purity forms of diamidino-2-phenylindole dye for fixed-cell, fluorescent staining of DNA content and nuclei for cellular imaging techniques.

Thermo Scientific Pierce DAPI Nuclear Counterstains are high-purity forms of diamidino-2-phenylindole dye for fixed-cell, fluorescent staining of DNA content and nuclei for cellular imaging techniques.

DAPI (diamidino-2-phenylindole) is a blue fluorescent probe that fluoresces brightly upon selectively binding to the minor groove of double stranded DNA, where its fluorescence is approximately 20-fold greater than in the nonbound state. Its selectivity for DNA and high cell permeability allows efficient staining of nuclei with little background from the cytoplasm. DAPI is a classic nuclear counterstain for immunofluorescence microscopy, as well as an important component of high-content screening methods requiring cell-based quantitation of DNA content.

Features of DAPI Nuclear Counterstain:

• DAPI dye—diamidino-2-phenylindole is a blue fluorescent stain specific for DNA
• Convenient—choose powder or easy-to-use DAPI dye solution (1 mg/mL)
• Cellular imaging—cell-permeable dye is effective for fixed-cell staining and quantitation of DNA content
• Counterstain—ideal for use alongside detection of specific targets with fluorescent antibodies for fluorescence microscopy or high-content screening (HCS)

Properties of DAPI Fluorescent Dye:

• Alternative names: DAPI Stain, DAPI Dye, DNA Content Counterstain
• Chemical name(s): 4,6'-diamidino-2-phenylindole, dihydrochloride; 4',6-diamidine-2-phenyl indole; 2-(4-Amidinophenyl)-6-indolecarbamidine dihydrochloride
• Molecular formula: C₁₆H₁₅N₅ · 2HCl
• Molecular weight: 350.25
• Excitation wavelength: 341 ± 3 nm (near 360 nm when bound to dsDNA)
• Emission wavelength: 452 ± 3 nm (456 to 460 nm when bound to dsDNA)
• Extinction coefficient: >30,600/M cm at 347 nm in methanol
• CAS #: 28718-90-3
• Purity: >95% (most lots >98%) by HPLC at 240 nm
• Solubility: >1 mg/mL in water; compound is soluble in DMF, water and various non-phosphate aqueous buffers
• Reactive groups: None; binds to minor groove of double-stranded DNA

As a counterstain in fluorescence imaging methods, DAPI is compatible with antibodies and other probes labeled with fluorescein (Cat. No. 53027) and rhodamine (Cat. No. 46112) dyes, as well as with Thermo Scientific DyLight Fluors. DAPI has greater photostability than Hoechst dyes, although Hoechst 33342 can be use for live cell imaging while use of DAPI is confined to fixed cells. DAPI is offered in powdered solid and aqueous solution forms.

Applications for DAPI Stain:

• Assaying DNA in solution (ref.4)
• Diagnosing mycoplasmal infection of cell cultures (ref.5)
• Measuring nuclear content and sorting chromosomes in flow cytometry (ref.6)
• Assessing apoptosis (ref.7)
• Detecting nuclei and organellar DNA in immunofluorescent and in situ hybridization procedures (ref.2,8)
• Replacing ethidium bromide for staining DNA in agarose gels (ref.5,9)
• Counterstaining nuclei in histochemical methods when red-fluorescent antibodies have been used to detect specific targets (ref.8)
• Reports also indicate that DAPI will bind to polyphosphates and other polyanions (ref.10), dextran sulfate (ref.11) and SDS (ref.12).

Related Products

• DAPI Solution, 1 mg/mL (Cat. No. 62248)

Specifications

• Color: Blue
• Content And Storage: Store at room temperature and protect from light.
• Excitation Wavelength Range: 360 nm
• Dye Type: Cell-Impermeant
• For Use With (Application): Cell imaging
• For Use With (Equipment): Fluorescence Microscope, High Content Instrument
• Quantity: 10 mg
• Detection Method: Fluorescence
• Description: DAPI (4',6-diamidino-2-phenylindole, dihydrochloride)
• Emission: 460 nm
• Product Line: Pierce
• Shipping Condition: Room Temperature
• Label Type: Fluorescent Dye
• Product Type: DAPI
• Sub Cellular Localization: Nucleus

Frequently Asked Questions (FAQs)

Is DAPI a good live-cell nuclear label?

DAPI is considered a semi-permeant/impermeant nucleic acid stain. Staining of nucleic is dependent upon the cell line in its performance. Some cell lines will label with DAPI, others not at all, and others label inconsistenly. Instead, we recommend using either Hoechst 33342 or Hoechst 33258, which have the same wavelength and binding mode as DAPI (at the A-T minor groove) but are readily cell-permeant.

I want to label the nuclei of live cells and track them over time. Can I use DAPI for this?

We do not recommend doing this. DAPI is considered to be a semi-permeant/impermeant nucleic acid stain. DAPI staining of live cells may be inconsistent. It is best used as a counterstain for fixed samples. Other cell permeable nucleic acid stains, such as Hoechst or the SYTO dyes may affect cellular function.

For mammalian cells, we recommend using the CellLight Nucleus transduction reagents, available in CFP, GFP and RFP. With these reagents, the cells are transduced overnight in a single labeling step and the next day the nuclei will fluoresce. The label may be retained for 3-5 days and should not affect cell function. Cytoplasmic cell tracking dyes such as the CellTracker dyes may also be used.

I notice that when I post-stain my cells with DAPI after performing the click reaction to detect EdU incorporation, my DAPI signal is lower compared to my no-click reaction control samples. What causes the reduction in DAPI signal?

The copper in the click reaction denatures DNA to a small extent (although not as much as is required for efficient BrdU detection), which can affect the binding affinity of DNA dyes including DAPI and Hoechst stain. This effect should only be apparent with the classic EdU kits and not the Click-iT Plus EdU kits, which use a lower copper concentration.

DAPI and Hoechst dyes are quite similar to each other. Why would I choose one over the other?

DAPI is a very common blue-fluorescent dye for nuclear counterstaining and gives very bright labeling on nuclei in fixed and permeabilized cells and tissues. However, it is considered to be a semi-permeant to impermeant stain and provides inconsistent staining of live cells. Hoechst 33342 dye is cell-permeant and stains with the same binding mechanism and fluorescent color; it is preferred for live-cell imaging and is just as good as DAPI for fixed cell labeling.

Safety and Handling

WARNING: Cancer - www.P65Warnings.ca.gov

For Research Use Only. Not for use in diagnostic procedures.