Portelite™ Fluorimetric Total Nucleic Acid Quantitation Kit Optimized for Cytocite™ and Qubit™ Fluorometers

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Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
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Shipping	Standard overnight for United States, inquire for international

Spectral properties

Excitation (nm)	509
Emission (nm)	527

Storage, safety and handling

H-phrase	H303, H313, H333
Hazard symbol	XN
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R22
UNSPSC	12171501

Overview SDS Protocol

Excitation (nm)

509

Emission (nm)

527

Portelite™ Fluorimetric Total Nucleic Acid Quantitation Kit is designed to rapidly measure the total amounts of nucleic acids, including double-stranded DNA (dsDNA), single-stranded DNA (ssDNA) and RNA in a sample. The kit has all the essential reagents, including Helixyte™ Green ssDNA reagent, dilution buffer, and prediluted DNA standards. Helixyte™ Green All reagent is a sensitive fluorescent nucleic acid probe for measuring the total amounts of nucleic acids in a sample that may contain double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), RNA and long oligonucleotides. Helixyte™ Green All reagent indiscriminately binds to dsDNA, ssDNA and RNA. Portelite™ Fluorimetric Total Nucleic Acid Quantitation Kit is optimized for measuring the total amounts of nucleic acids with CytoCite™ or Qubit® fluorometers.

Platform

Qubit Fluorometer

Excitation	480 nm
Emission	530 nm
Instrument specification(s)	0.2 mL PCR tube

CytoCite Fluorometer

Excitation	480 nm
Emission	530 nm
Instrument specification(s)	0.2 mL PCR tube

Components

Example protocol

AT A GLANCE

Protocol Summary

Prepare a Helixyte™ Green All working solution
Add 190 µL of 1X Helixyte™ Green All working solution into each 0.2 mL PCR tube
Add 10 µL of Nucleic Acid Standards or test samples into each tube
Incubate at room temperature for 2 minutes
Monitor the fluorescence intensity with CytoCite™ fluorometer or Qubit™ fluorometer

Important

All kit components must be brought to room temperature before starting the experiment.

PREPARATION OF WORKING SOLUTION

Helixyte™ Green All working solution

Make a 200-fold dilution of Helixyte™ Green All reagent (Component A) with Assay Buffer (Component B). For example, to prepare enough working solution for 5 samples, add 5 μL of Helixyte™ Green All (Component A) into 1 mL of Assay Buffer (Component B).
Note Protect the working solution from light by covering it with foil or placing it in the dark. It’s recommended to prepare the solution in a plastic container rather than a glass container, as the dye may adsorb to the glass surface. For best results, this solution should be used within a few hours after the dilution.

SAMPLE EXPERIMENTAL PROTOCOL

The acceptable range for the sample volume could be 1~20 μL depending on the estimated concentration of the Nucleic Acid sample.
The following protocol is generated based on a sample volume of 10 μL.

Add 190 µL of 1X Helixyte™ Green All working solution into each Cytocite™ sample tube (#CCT100) or the equivalent 0.2 mL PCR tube.
Note Use thin-wall, polypropylene, clear 0.2 mL PCR tubes such as AAT Cat#CCT100.
Add 10 μL of Nucleic Acid Standards or test samples into each tube, and then mix by vortexing for 2~3 seconds.
Incubate all tubes at room temperature for 2 minutes.
Insert the samples into CytoCite™ or Qubit™ and monitor the fluorescence intensity with the green fluorescence channel. Follow the appropriate procedures for CytoCite™ Fluorometer. See the link below for detailed instructions: https://devices.aatbio.com/documentation/user-manual-for-cytocite-fluorometer

Preparation of Standard Calibration Curve

For Portelite™ assays, you have the choice to make a calibration curve with the Nucleic Acid Standards. Here is a brief protocol to generate a customized DNA standard curve.

Perform a 1:2 serial dilution: Add 10 ng/μL Nucleic Acid Standard #2 (Component D) into the Assay Buffer (Component B) to get 10, 5, 2.5, 1.25, 0.62, 0.31, 0.15 ng/µL DNA standard dilutions.
Add 190 µL of the Helixyte™ Green All working solution into each tube.
Add 10 µL of standards into a 0.2 mL PCR tube and then mix by vortexing for 2∼3 seconds.
Incubate the reaction at room temperature for 2 minutes.
Insert the samples into CytoCite™ and monitor the fluorescence intensity with the green fluorescence channel.

Spectrum

Open in Advanced Spectrum Viewer

Spectral properties

Excitation (nm)	509
Emission (nm)	527

Images

Figure 1. Comparison of total nucleic acid dose response using the Qubit™ fluorometer (blue) or CytoCite™ fluorometer (red).

References

View all 13 references: Citation Explorer

Accurate bulk quantitation of droplet digital PCR.
Authors: Sun, Chen and Liu, Leqian and Vasudevan, Harish N and Chang, Kai-Chun and Abate, Adam R
Journal: bioRxiv : the preprint server for biology (2021)

Nucleic Acid Quantitation with Log-Linear Response Hybridization Probe Sets.
Authors: Wu, Lucia R and Fang, John Z and Khodakov, Dmitriy and Zhang, David Yu
Journal: ACS sensors (2020): 1604-1614

Nucleic Acid Extraction from Human Biological Samples.
Authors: Mullegama, Sureni V and Alberti, Michael O and Au, Cora and Li, Yan and Toy, Traci and Tomasian, Vanina and Xian, Rena R
Journal: Methods in molecular biology (Clifton, N.J.) (2019): 359-383

MICROFLUIDIC DEVICES FOR LABEL-FREE AND NON-INSTRUMENTED QUANTITATION OF UNAMPLIFIED NUCLEIC ACIDS BY FLOW DISTANCE MEASUREMENT.
Authors: Chatterjee, Debolina and Mansfield, Danielle S and Woolley, Adam T
Journal: Analytical methods : advancing methods and applications (2014): 8173-8179

Helicase-dependent amplification of nucleic acids.
Authors: Cao, Yun and Kim, Hyun-Jin and Li, Ying and Kong, Huimin and Lemieux, Bertrand
Journal: Current protocols in molecular biology (2013): 15.11.1-15.11.12

Microvolume quantitation of nucleic acids.
Authors: Desjardins, Philippe R and Conklin, Deborah S
Journal: Current protocols in molecular biology (2011): 3J

Concentration determination of nucleic acids and proteins using the micro-volume BioSpec-nano-spectrophotometer.
Authors: Sukumaran, Suja
Journal: Journal of visualized experiments : JoVE (2011)

NanoDrop microvolume quantitation of nucleic acids.
Authors: Desjardins, Philippe and Conklin, Deborah
Journal: Journal of visualized experiments : JoVE (2010)

Comparison of two real-time PCR methods for detection of ostreid herpesvirus 1 in the Pacific oyster Crassostrea gigas.
Authors: Martenot, C and Oden, E and Travaillé, E and Malas, J P and Houssin, M
Journal: Journal of virological methods (2010): 86-9

Effect of perinatal short-course zidovudine on the clinical and virological manifestations of HIV-1 subtype E infection in infants.
Authors: Sutthent, Ruengpung and Chokephaibulkit, Kulkanya and Piyasujabul, Daorung and Vanprapa, Nirun and Roogpisuthipong, Anuwat and Chaisilwatana, Pongsakdi
Journal: Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology (2002): 47-56