Method of quantitating dsDNA
- Los Alamos, NM
A method for quantitating dsDNA in an aqueous sample solution containing an unknown amount of dsDNA. A first aqueous test solution containing a known amount of a fluorescent dye-dsDNA complex and at least one fluorescence-attenutating contaminant is prepared. The fluorescence intensity of the test solution is measured. The first test solution is diluted by a known amount to provide a second test solution having a known concentration of dsDNA. The fluorescence intensity of the second test solution is measured. Additional diluted test solutions are similarly prepared until a sufficiently dilute test solution having a known amount of dsDNA is prepared that has a fluorescence intensity that is not attenuated upon further dilution. The value of the maximum absorbance of this solution between 200-900 nanometers (nm), referred to herein as the threshold absorbance, is measured. A sample solution having an unknown amount of dsDNA and an absorbance identical to that of the sufficiently dilute test solution at the same chosen wavelength is prepared. Dye is then added to the sample solution to form the fluorescent dye-dsDNA-complex, after which the fluorescence intensity of the sample solution is measured and the quantity of dsDNA in the sample solution is determined. Once the threshold absorbance of a sample solution obtained from a particular environment has been determined, any similarly prepared sample solution taken from a similar environment and having the same value for the threshold absorbance can be quantified for dsDNA by adding a large excess of dye to the sample solution and measuring its fluorescence intensity.
- Research Organization:
- Los Alamos National Laboratory (LANL), Los Alamos, NM (United States)
- DOE Contract Number:
- W-7405-ENG-36
- Assignee:
- The Regents of the University of California (Los Alamos, NM)
- Patent Number(s):
- US 6350578
- OSTI ID:
- 874251
- Country of Publication:
- United States
- Language:
- English
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quantitating
dsdna
aqueous
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dye-dsdna
complex
fluorescence-attenutating
contaminant
prepared
fluorescence
intensity
measured
diluted
provide
concentration
additional
solutions
similarly
sufficiently
dilute
attenuated
dilution
value
maximum
absorbance
200-900
nanometers
nm
threshold
identical
chosen
wavelength
dye
added
form
dye-dsdna-complex
quantity
determined
obtained
environment
similar
quantified
adding
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