The assay kit utilizes Picogreen DNA dye, which specifically binds to double-stranded DNA without binding to proteins, RNA, or salt ions. By exciting the sample with light and collecting the emitted light using a Qubit fluorometer, the actual content of double-stranded DNA in the sample can be accurately quantified. This method eliminates interference from substances such as proteins, RNA, and salt ions, providing precise quantification of double-stranded DNA.

        Nanodrop series microvolume spectrophotometers use the commonly used UV absorption method to detect the concentration and purity of the samples. However, because they measure the absorbance of all substances at 260nm (such as DNA, RNA, degraded nucleic acids, and free nucleotides), the readings are not always accurate. In contrast, fluorescence assays measure the concentration of target molecules by detecting the fluorescence intensity after specific binding of fluorescent dyes. Therefore, their quantification results are generally lower than the readings at A260nm but are more accurate.

        Moreover, accurate DNA quantification is crucial for many downstream applications. Nanodrop series full-wavelength microvolume spectrophotometers cannot accurately measure DNA concentrations below 5ng/µl, which many DNA samples fall within. In contrast, the detection limit of a fluorescence assay for double-stranded DNA can be as low as 0.5pg/µl. Therefore, a fluorescence assay and the corresponding quantification reagents offer a faster, more sensitive, and more accurate method for determining DNA concentrations.

        This assay will be used for costly downstream experiments such as qPCR, PCR cloning, transfection, and next-generation sequencing, where precise quantification in the range of 10 pg/µL to 100 ng/µL is essential.

Comparative results between Thermofisher (Cat.no.Q32854) test standards and samples are shown in Figure 1.

Figure 2 demonstrates the stability of the test standards and samples.

        Testing was performed using both Q32854 and iPure HS dsDNA assay kit for the standards and three samples. The results showed high consistency, indicating that the iPure HS dsDNA assay kit can fully replace Q32854 for testing. Stability testing of the iPure HS dsDNA assay kit stored at room temperature, 4°C, and 37°C for 1 day, 3 days, 7 days, and 1 month showed no significant differences in sample measurements, indicating good kit stability.