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Helixyte™ Green Fluorimetric dsDNA Quantitation Kit *High Sensitivity*

Comparison of dsDNA dose response using the Helixyte Green™ (blue) with Invitrogen<sup>TM</sup> Quant-iT™ PicoGreen® dsDNA Reagent (red ). dsNDA standards were incubated in cuvettes and measured using varian cary eclipse fluorescence spectrophotometer. 
Comparison of dsDNA dose response using the Helixyte Green™ (blue) with Invitrogen<sup>TM</sup> Quant-iT™ PicoGreen® dsDNA Reagent (red ). dsNDA standards were incubated in cuvettes and measured using varian cary eclipse fluorescence spectrophotometer. 
Ordering information
Price ()
Catalog Number17651
Unit Size
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Additional ordering information
Telephone1-408-733-1055
Fax1-408-733-1304
Emailsales@aatbio.com
InternationalSee distributors
ShippingStandard overnight for United States, inquire for international
Spectral properties
Excitation (nm)502
Emission (nm)522
Storage, safety and handling
H-phraseH303, H313, H340
Hazard symbolT
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R68
UNSPSC41116134

OverviewpdfSDSpdfProtocol


Excitation (nm)
502
Emission (nm)
522
Helixyte™ Green dsDNA Quantitation Assay Kit can be used for selectively detecting as little as 25 pg/ml of dsDNA in the presence of ssDNA, RNA, and free nucleotides. Helixyte™ Green exhibits large fluorescence enhancement upon binding to dsDNA. The assay is linear over three orders of magnitude and has little sequence dependence, allowing you to accurately measure DNA from many sources, including genomic DNA, viral DNA, miniprep DNA, or PCR amplification products. Helixyte™ Green dsDNA Quantitation Assay Kit is a few magnitudes more sensitive than UV absorbance readings. It is specific for dsDNA in the presence of equimolar amounts of RNA. The kit is robust with a mix and read format. It can be used with a bench top fluorometer or a hand-held fluorescence meter (e.g., Qubit fluorometer). This kit is an excellent replacement for Quant-iT™ PicoGreen® dsDNA Assay Kit (Quant-iT™ and PicoGreen® are the trademarks of Invitrogen).

Platform


Spectrofluorometer

Excitation490 nm
Emission525 nm
Cutoff515 nm

Components


Component A: Helixyte Green™1 vial (1 mL, 200X in DMSO)
Component B: 20X Assay buffer1 bottle (25 mL)
Component C: Calf thymus DNA Standard1 vial (1 mL, 100 µg/mL)

Example protocol


AT A GLANCE

Protocol summary

  1. Add 1mL dsDNA standards or test samples to each cuvette
  2. Add 1mL Helixyte Green™ working solution 
  3. Incubate at RT for 5-10 minutes
  4. Monitor the fluorescence at Ex/Em=490/525 nm

Important notes
The following protocol is an example for quantifying dsDNA with Helixyte Green™. Allow all the components to warm to room temperature before opening. No data is available addressing the mutagenicity or toxicity of Helixyte Green™dsDNA stain. Because this reagent binds to nucleic acids, it should be treated as a potential mutagen and handled with appropriate care. The DMSO stock solution should be handled with particular caution as DMSO is known to facilitate the entry of organic molecules into tissues.

PREPARATION OF STOCK SOLUTION

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.

1. Assay Buffer (1X)
Prepare a 1X Assay buffer by diluting the concentrated buffer 20-fold with sterile, distilled, DNase-free water.

PREPARATION OF STANDARD SOLUTION

dsDNA standard

For convenience, use the Serial Dilution Planner: https://www.aatbio.com/tools/serial-dilution/17651

For high range standard curve:
Add 30 µL of 100 µg/mL dsDNA stock solution (Component C) to 1.47 mL of 1X Assay buffer to have 2000 ng/mL dsDNA solution, and then perform 1:2 and 1:10 serial dilutions to get 1000, 100, 10, 1 and 0 ng/mL.

For low range standard curve:
Add 40 µL of 2 µg/mL dsDNA stock solution to 1.56 mL of 1X Assay buffer to have 50 ng/mL dsDNA solution, and then perform 1:2 and 1:10 serial dilutions to get 25, 2.5, 0.25, 0.025 and 0 ng/mL.

PREPARATION OF WORKING SOLUTION

Prepare Helixyte Green™ working solution by making a 200-fold dilution of the concentrated DMSO solution in 1X assay buffer. For example, to prepare enough working solution to assay 10 samples in a 2 mL final volume, add 50 μL of Helixyte Green™ (Component A) into 10 mL of Assay Buffer (Component B). Protect the working solution from light by covering it with foil or placing it in the dark. Note: We recommend preparing this solution in a plastic container rather than glass, as the dye may adsorb to glass surfaces. For best results, this solution should be used within a few hours of its preparation.

SAMPLE EXPERIMENTAL PROTOCOL

  1. Add 1 mL of Helixyte Green™ working solution to each cuvette containing 1 mL of the dsDNA standard, blank control, and test samples to make the total dsDNA assay volume of 2 mL/cuvette.

  2. Incubate the reaction at room temperature for 5 to 10 minutes, protected from light.

  3. Monitor the fluorescence increase with a spectroflurometer at Ex/Em = 490/525 nm. Note: To minimize photobleaching effects, keep the time for fluorescence measurement constant for all samples.

     

Spectrum


Open in Advanced Spectrum Viewer
spectrum

Spectral properties

Excitation (nm)502
Emission (nm)522

References


View all 31 references: Citation Explorer
Inhibitors of Streptococcus pneumoniae surface endonuclease EndA discovered by high-throughput screening using a PicoGreen fluorescence assay
Authors: Peterson EJ, Kireev D, Moon AF, Midon M, Janzen WP, Pingoud A, Pedersen LC, Singleton SF.
Journal: J Biomol Screen (2013): 247
Validation of a PicoGreen-based DNA quantification integrated in an RNA extraction method for two-dimensional and three-dimensional cell cultures
Authors: Chen Y, Sonnaert M, Roberts SJ, Luyten FP, Schrooten J.
Journal: Tissue Eng Part C Methods (2012): 444
Characterization of PicoGreen interaction with dsDNA and the origin of its fluorescence enhancement upon binding
Authors: Dragan AI, Casas-Finet JR, Bishop ES, Strouse RJ, Schenerman MA, Geddes CD.
Journal: Biophys J (2010): 3010
Comparison of SYBR Green I-, PicoGreen-, and [3H]-hypoxanthine-based assays for in vitro antimalarial screening of plants from Nigerian ethnomedicine
Authors: Abiodun OO, Gbotosho GO, Ajaiyeoba EO, Happi CT, Hofer S, Wittlin S, Sowunmi A, Brun R, Oduola AM.
Journal: Parasitol Res (2010): 933
Metal-enhanced PicoGreen fluorescence: application to fast and ultra-sensitive pg/ml DNA quantitation
Authors: Dragan AI, Bishop ES, Casas-Finet JR, Strouse RJ, Schenerman MA, Geddes CD.
Journal: J Immunol Methods (2010): 95
Quantification of dsDNA using the Hitachi F-7000 Fluorescence Spectrophotometer and PicoGreen dye
Authors: Moreno LA, Cox KL.
Journal: J Vis Exp. (2010)
Factors affecting quantification of total DNA by UV spectroscopy and PicoGreen fluorescence
Authors: Holden MJ, Haynes RJ, Rabb SA, Satija N, Yang K, Blasic JR, Jr.
Journal: J Agric Food Chem (2009): 7221
Development and characterization of a novel host cell DNA assay using ultra-sensitive fluorescent nucleic acid stain "PicoGreen"
Authors: Ikeda Y, Iwakiri S, Yoshimori T.
Journal: J Pharm Biomed Anal (2009): 997
Enhanced DNA dynamics due to cationic reagents, topological states of dsDNA and high mobility group box 1 as probed by PicoGreen
Authors: Noothi SK, Kombrabail M, Kundu TK, Krishnamoorthy G, Rao BJ.
Journal: FEBS J (2009): 541
Label-free DNA sequence detection with enhanced sensitivity and selectivity using cationic conjugated polymers and PicoGreen
Authors: Ren X, Xu QH.
Journal: Langmuir (2009): 43