PicoGreen is a high-sensitivity and high-specificity double-stranded DNA (dsDNA) stain. It binds specifically to dsDNA and emits green fluorescence, and its fluorescence intensity is proportional to the amount of dsDNA in the sample. Therefore, dsDNA in samples can be quantified by measuring fluorescence readouts. This stain delivers ultrahigh sensitivity for dsDNA detection, with a limit of detection down to the pg/mL level. In addition, compared with the traditional UV absorbance method for DNA quantification, PicoGreen exhibits excellent specificity; it is barely interfered by single-stranded DNA (ssDNA), proteins and salts in samples, which greatly improves quantification accuracy. This stain is applicable to a wide range of molecular biology experiments, including accurate quantification and normalization of NGS libraries; DNA sample quality control prior to qPCR or microarray analysis; viral vector titer determination and evaluation of cellular genomic DNA extraction yield; quantification of trace nucleic acids such as environmental microbial DNA and forensic samples.
Product Advantages
1. Ultrahigh sensitivity: Detects dsDNA at concentrations as low as 25 pg/mL;
2. Excellent specificity: Shows negligible response to RNA, single-stranded DNA or free nucleotides, resulting in low background interference;
3. Strong fluorescence signal: Fluorescence signal enhancement of approximately 1,000-fold upon binding to dsDNA;
4. Wide linear dynamic range: Quantifies dsDNA across 4 orders of magnitude;
5. Simple and rapid workflow: Simply add the stain to samples and incubate before detection;
6. Safe and eco-friendly: PicoGreen has low toxicity and ensures safe handling.
Operating Procedures
Reagents Required Before Experiment: 1× TE Buffer, Standard DNA
I. dsDNA Quantification with Qubit® Fluorometer
Take a 10 μL dsDNA sample as an example:
1. Equilibrate PicoGreen stain to room temperature, fully dissolve it, vortex thoroughly and perform a brief centrifugation.
2. Dilute the stain 200-fold with 1× TE Buffer to prepare working stain solution.
3. Add 190 μL working solution into Qubit assay tubes according to the number of test samples.
4. Add 10 μL of dsDNA Standard 1, dsDNA Standard 2 or unknown dsDNA sample to each tube, vortex for 2–3 seconds, and incubate at room temperature in the dark for 1 min.
5. Follow the operating instructions of the Qubit® fluorometer, select the dsDNA High Sensitivity assay program to measure fluorescence signal values.
II. dsDNA Quantification with Microplate Reader
Take a 10 μL dsDNA sample as an example:
1. Equilibrate PicoGreen stain to room temperature, fully dissolve it, vortex thoroughly and perform a brief centrifugation.
2. Dilute the stain 200-fold with 1× TE Buffer to prepare working stain solution.
3. Add 190 μL working solution into a 96-well microplate based on the number of samples to be tested.
4. Prepare standard DNA with 5 distinct concentration gradients; the dsDNA concentration of unknown samples should fall within the range of these 5 gradients.
5. Load unknown dsDNA samples and standard concentration samples into the 96-well microplate, then incubate at room temperature for 2 min.
6. Measure fluorescence signals via the microplate reader, set excitation wavelength to 480 nm and emission wavelength to 520 nm.
7. Generate a standard curve based on standard concentrations and corresponding fluorescence signals, then calculate the concentration of unknown samples using the standard curve and sample fluorescence values.
Precautions
1. Wear a lab coat and disposable gloves during operation for personal safety and health protection.
2. This product is for research use only.
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