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Helix NP™ Green

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RUO
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Helix_NP_Green_1_FC_112916
One-day-old C57BL/6 mouse splenocytes were stained with 5 µM Helix NP™ Green and APC Annexin V with no wash.
  • Helix_NP_Green_1_FC_112916
    One-day-old C57BL/6 mouse splenocytes were stained with 5 µM Helix NP™ Green and APC Annexin V with no wash.
  • Helix_NP_Green_ICC_012821
    HeLa cells were fixed with 1% paraformaldehyde (PFA) for ten minutes, permeabilized with 0.5% Triton X-100 for ten minutes, and blocked with 5% FBS for 30 minutes. Then the cells were intracellularly stained with 5 µg/mL of Alexa Fluor® 647 anti-Cytokeratin (pan reactive) antibody (red) in blocking buffer overnight followed by 1:500 dilution of Helix NP™ Green (green) and Flash Phalloidin™ Red 594 (blue) staining for 15 minutes at room temperature. The image was captured with a 60X objective.
  • Helix_NP_Green_IHC-F_012821
    Mouse frozen cerebellum tissue was fixed with 4% paraformaldehyde (PFA) for ten minutes, permeabilized with 0.5% Triton X-100 for ten minutes, and blocked with 5% FBS for one hour. Then the tissue was intracellularly stained with 1:500 dilution of Helix NP™ Green (green) for 15 minutes at room temperature. The image was captured with a 20X objective.
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425303 250 µL $211
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Description

Helix NP™ Green is a green-emitting nucleic acid stain. It is impermeant to live cells and thus can be used for the discrimination of live and dead cells. In immunofluorescence microscopy, it can be used as a nuclear counterstain in cells and tissue. It is optimally excited at 495 nm with an emission at 519 nm, which can be detected in the Alexa Fluor® 488 or FITC channel.

Product Details
Technical data sheet

Product Details

Preparation
Helix NP™ Green is supplied at 250 µl per vial.
Concentration
5.0 mM
Storage & Handling
Upon receipt, store at -20°C.
Application

FC - Quality tested
IF, IHC-F - Verified
Recommended Usage

For use in viability assays, the optimal concentration can range from 1 - 10 nM. For immunofluorescence microscopy and immunohistochemical staining on frozen tissue sections, the suggested use of this reagent is 0.5 - 10 µM. It is recommended that the reagent be titrated for optimal performance for each application.

Application Notes

Helix NP™ Green is a cell-impermeant nucleic acid probe suitable for use as a viability dye in flow cytometry. It can also be used for viability in microscopy on live cells or as a nuclear counterstain on fixed and permeabilized cells and tissues. It is a green-emitting dye with an excitation/emission max of 495 nm/519 nm that can be detected in the Alexa Fluor® 488 or FITC channel.

Protocol for flow cytometric viability staining using Helix NP™ Green:

  1. Isolate cells following protocol of choice.
  2. Optional— For multicolor flow cytometry experiments, surface-stain cells as recommended. Helix NP Green should be added as the last step prior to sample acquisition.
    • Note: The use of Helix NP for viability staining is incompatible with cell fixation and permeabilization protocols. If cells are to be fixed and permeabilized, use a fixable viability dye, such as a Zombie™ Fixable Viability kit.
  3. Dilute Helix NP Green to required concentration. We have observed good flow cytometric viability staining in a final concentration of 1 - 10 nM, but recommend titrating the reagent to determine optimal concentration for cells of interest.
    • For example, to stain cells in a final concentration of 10 nM Helix NP Green, prepare a 1:5000 dilution of the 5.0 mM stock in Cell Staining buffer (or equivalent). Then, add 5 µL of diluted reagent to 500 µL of cell suspension.
  4. Do not wash the cells after adding Helix NP™ Green. Samples are ready for acquisition.
  5. Analyze cells on a cytometer equipped with a 488 nm blue laser.

Protocol for nuclear counterstaining fixed and permeabilized cell /tissue specimens using Helix NP™ Green:

  1. Fix specimens with 1%-4% Paraformaldehyde (PFA) for 10 minutes at room temperature.
    • 1% for cultured cells
    • 4% for frozen tissue
  2. Wash the cells/tissue two times with 1X PBS.
  3. Permeabilize the cells/tissue with 0.5% Triton X-100 for 10 minutes at room temperature.
  4. Wash the cells/tissue two times with 1X PBS.
  5. Block cells with 5% fetal bovine serum for 30 minutes at room temperature.
  6. Complete any additional blocking steps, antibody staining, and washes before adding the Helix NP™ Green counterstain.
  7. Prepare the working solution.
    • It is best to try a range of dye concentrations to determine the optimal concentration for cells/tissues of interest to achieve the best image. We have obtained good results using a range of 0.5 - 10 µM for IHC-F and IF/ICC).
  8. Stain the cells/tissues with the working solution for 20 minutes at 4°C or room temperature in the dark.
    • Protect from light prior to imaging.
    • No wash step is needed after staining.
  9. Mount the slides with an antifade medium.
  10. Image slides with a filter ideal for ex/em max of 495 nm/519 nm (Alexa Fluor® 488 or FITC channel.

Antigen Details

Biology Area
Apoptosis/Tumor Suppressors/Cell Death, Cell Biology, Cell Cycle/DNA Replication, Cell Proliferation and Viability
Gene ID
NA
Go To Top Version: 6    Revision Date: 05/30/2024

For Research Use Only. Not for diagnostic or therapeutic use.

 

This product is supplied subject to the terms and conditions, including the limited license, located at www.biolegend.com/terms) ("Terms") and may be used only as provided in the Terms. Without limiting the foregoing, BioLegend products may not be used for any Commercial Purpose as defined in the Terms, resold in any form, used in manufacturing, or reverse engineered, sequenced, or otherwise studied or used to learn its design or composition without express written approval of BioLegend. Regardless of the information given in this document, user is solely responsible for determining any license requirements necessary for user’s intended use and assumes all risk and liability arising from use of the product. BioLegend is not responsible for patent infringement or any other risks or liabilities whatsoever resulting from the use of its products.

 

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