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QUBUIT DNA QUANTIFICATION SOFTWARE

How many samples can be measured at a time? (throughput) What is the upper detection limit (dsDNA)?ĭepends on sample format and volume used (in the case of microplates)

What is the lower detection limit (dsDNA)?ĭepends on sample format and volume used (in the case of microplates)* It takes longer-reagent and sample preparation are required.It has limited sensitivity-detection limits are higher than fluorescence-based methods.

QUBUIT DNA QUANTIFICATION SOFTWARE

It is not selective-uses software algorithms to distinguish mammalian DNA and RNA.It is accurate despite contamination being present in the sample, including nucleic acid contaminants.It is sensitive-can measure pg/mL it is the recommended method for very diluted nucleic acid samples.It is specific-performed measurement is selective for DNA or RNA.Can provide information on contaminants-can identify non-nucleic acid contamination in samples (proteins, phenol, guanidine salts) as well as mammalian DNA/RNA contamination and provide corrected concentrations (applicable to NanoDrop One/One C/Eight instruments).Can provide direct measurements of purity ratios-A260/280 and A260/230.It is simple-no sample preparation, dyes, or standards are required.The limit of detection and linear response of the measurements are specific to each assay.Concentrations of nucleic acids are measured using the fluorescence signal of the sample and a calibration curve is generated from standard samples of known concentration and fit to appropriate regression models.This concentration calculation is automated in many instruments.ε=wavelength-dependent molar absorptivity coefficient (or extinction coefficient) in M -1cm -1.

A=UV absorbance in absorbance units (AU).

C= nucleic acid concentration in molar (M).

Concentrations of nucleic acids can be directly calculated from their measured absorbance values at 260 nm, using the Beer-Lambert's equation:.

How is the concentration of nucleic acids calculated?

Wavelength separation can take place in various ways (for example with filters or with monochromators).

Wavelength separation can take place before or after the light has passed the sample, and the optical light path can be horizontal or vertical.

Sample is excited with filtered light (at the excitation wavelength, and the emitted light (at the emission wavelength) is recorded by a detector.

The signal is measured by fluorometers.

The attenuation in the light that reaches the detector after passing through the sample is measured in relation to the incident light and expressed as absorbance values of the sample in the solution.

The signal is measured by spectrophotometers or spectrometers.

Dyes only emit signal when bound to the target. The fluorometric measurement of nucleic acids is based upon the use of fluorogenic dyes that bind selectively to DNA or RNA.įluorescent dyes selectively bind to DNA, RNA or protein. When an absorption spectrum is measured, nucleic acids absorb light with a characteristic peak at 260 nm. The photometric measurement of nucleic acids is based on the intrinsic absorptivity properties of nucleic acids (DNA and RNA).