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US 6,999,173 B2Granted
Method and apparatus for ratio fluorometry
AssigneeFFA Sciences LLC
InventorsAlan M. Kleinfeld, Pavel V. Vodkin
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Abstract
A method and apparatus for determining the concentration of molecules or atoms by measuring the ratio of two fluorescence signals. A sample having fluorescent molecules is exposed to radiation or excitation energy from a first source, which can be a broadband light source. The fluorescence of the sample is detected at two different wavelengths. The concentration of specific molecules or atoms within the sample is determined using the ratio of the two fluorescence signals. Fluorescent molecules can be bound to a human serum or plasma sample to allow determination of the concentration of unbound free fatty acids in the sample.
Claims80
An apparatus comprising:
- a sample receptacle;
- an excitation source providing an excitation emission directed towards the sample receptacle;
- a first detector configured to detect a first sample emission;
- a second detector configured to detect a second sample emission, wherein an axis of the first sample emission is the same as an axis of the excitation emission;
- and
- a processor configured to calculate a ratio of the detected first sample emission to the detected second sample emission.
3 more elements
A ratio fluorometer for determining a concentration of a target molecule in a sample, the fluorometer comprising:
- a sample receptacle configured to support the sample;
- an excitation light source directed towards the sample receptacle and configured to generate an excitation emission, wherein an axis of the excitation emission is the same as an axis of a first fluorescence from the sample;
- a first optical filter configured to filter the first fluorescence from the sample;
- a first detector configured to detect an output of the first optical filter;
- a first Analog to Digital Converter (ADC) configured to generate a digital representation of an output of the first detector;
- a second optical filter configured to filter a second fluorescence from the sample;
- a second detector configured to detect an output from the second optical filter;
- a second ADC configured to generate a digital representation of an output of the second detector;
- and
- a processor configured to calculate a ratio, based in part, on the digital representations of the output of the first detector and the output of the second detector and to determine a concentration of the target molecule based, at least in part, on the ratio.
7 more elements
A ratio fluorometer for determining a concentration of unbound free fatty acids (FFAu) in a sample, the fluorometer comprising:
- a sample receptacle configured to support the sample;
- an excitation source configured to generate an excitation emission, wherein an axis of the excitation emission is the same as an axis of a first sample emission;
- a first detector configured to detect a first fluorescence of a molecule when the molecule is bound to a ligand of the free fatty acid in the sample;
- a second detector configured to detect a second fluorescence of the molecule when the molecule is unbound to the ligand of the free fatty acid in the sample;
- and
- a processor configured to calculate a ratio, based at least in part, on an output of the first detector and an output of the second detector, and based at least in part on the ratio, to determine the FFAu concentration in the sample.
3 more elements
A method comprising:
- exciting a sample with an excitation emission;
- detecting a first emission from the sample to produce a first detected emission, wherein an axis of the first sample emission is the same as an axis of the excitation emission;
- detecting a second emission from the sample to produce a second detected emission;
- and
- determining a concentration of a molecule in the sample based at least in part on the ratio of the detected emissions.
2 more elements
A method of determining a concentration of a target molecule in a sample, the method comprising:
- exciting the sample with an excitation emission from an excitation light source;
- detecting, using a first detector, a first fluorescence from the sample, wherein an axis of the first fluorescence is the same as an axis of the excitation emission;
- converting an output of the first detector to a first digital representation;
- detecting, using a second detector, a second fluorescence from the sample;
- converting an output of the second detector to a second digital representation;
- calculating a ratio based, at least in part, on the first digital representation and the second digital representation;
- and
- determining the concentration of the target molecule based, at least in part, on the ratio.
5 more elements
A method of determining a concentration of unbound free fatty acids (FFAu) in a sample, the method comprising:
- determining a fluorescence of bound molecules from the sample using a first detector and epifluorescence;
- determining a fluorescence of unbound molecules from the sample using a second detector;
- and
- determining the concentration of FFAu in the sample based, at least in part, on a ratio of the fluorescence of bound molecules to the fluorescence of unbound molecules.
1 more element
A measurement apparatus comprising:
- a means for receiving a sample;
- a means for exciting the sample;
- a first means for detecting first sample emissions and configured with epifluorescence;
- a first means for generating a first digital output corresponding to an output from the first means for detecting;
- a second means for detecting second sample emissions;
- a second means for generating a second digital output corresponding to an output from the second means for detecting;
- and
- a processor configured to determine a concentration of target molecules in the sample based, at least in part, on a ratio of the first digital output to the second digital output.
5 more elements
A measurement apparatus, comprising:
- a first means for filtering a sample emission to produce a first filtered emissions, wherein the measurement apparatus is configured with epifluorescence;
- a first means for detecting the first filtered emission and producing a first detected emission value;
- a second means for filtering the sample emission to produce a second filtered emission;
- a second means for detecting the second filtered emission and producing a second detected emission value;
- and
- a processor coupled to the first means for detecting and the second means for detecting and configured to determine, based in part on a ratio of the first detected emission value to the second detected emission value, a concentration value.
3 more elements
A method of determining a concentration of a target molecule in a sample, the method comprising:
- detecting a first fluorescence from the sample in a first bandwidth to produce a first detected value, wherein the first fluorescence is detected using epifluorescence;
- detecting a second fluorescence from the sample in a second bandwidth to produce a second detected value;
- calculating a ratio based, at least in part, on the first detected value and the second detected value;
- and
- determining the concentration of the target molecule based, at least in part, on the ratio.
2 more elements
A method of calibrating a ratio fluorometer;
- the method comprising:
- determining in a calibrating ratio fluorometer a ratio of fluorescence of a sample;
- determining in a second ratio fluorometer an uncorrected ratio of fluorescence of the sample;
- and
- adjusting a detector output such that the uncorrected ratio of fluorescence is substantially equal to the ratio of fluorescence.
2 more elements
Dependencies are inferred from the claim text.