Waveguide-based detection system with scanning light source
The invention provides methods and devices for generating optical pulses in one or more waveguides using a spatially scanning light source. A detection system, methods of use thereof and kits for detecting a biologically active analyte molecule are also provided. The system includes a scanning light source, a substrate comprising a plurality of waveguides and a plurality of optical sensing sites in optical communication with one or more waveguide of the substrate, a detector that is coupled to and in optical communication with the substrate, and means for spatially translating a light beam emitted from said scanning light source such that the light beam is coupled to and in optical communication with the waveguides of the substrate at some point along its scanning path. The use of a scanning light source allows the coupling of light into the waveguides of the substrate in a simple and cost-effective manner.
1. A method of measuring a concentration of an analyte using a waveguide chip, the method comprising:
adding a known amount of one or more normalizing moieties to a sample;
adding a first detection molecule labelled with a first fluorescent tag to the sample, the first detection molecule specific to the analyte;
adding one or more secondary detection molecules, each secondary detection molecule labelled with a distinct secondary fluorescent tag, to the sample, the one or more secondary detection molecules specific to the one or more normalizing moieties, wherein the first fluorescent tag and the one or more secondary fluorescent tags emit at different fluorescence wavelengths;
introducing the sample to an array of sensing sites on the waveguide chip, each sensing site of the array having a first capture molecule specific to the analyte and one or more secondary capture molecules specific to the one or more normalizing moieties, wherein the first capture molecule and the one or more secondary capture molecules are both immobilized in each sensing site of the array at a predetermined ratio;
introducing a first light pulse into one or more excitation waveguides on the waveguide chip at a first predetermined interval, the one or more excitation waveguides in optical communication with the array of sensing sites, the first light pulse configured to excite the first fluorescent tag;
introducing one or more secondary light pulses into the one or more excitation waveguides on the waveguide chip at one or more secondary predetermined intervals, wherein the first light pulse and the one or more secondary light pulses have different wavelengths, the one or more secondary light pulses configured to excite the one or more secondary fluorescent tags;
measuring the fluorescence emitted by the first detection molecule and the one or more secondary detection molecules through a plurality of collection waveguides in optical communication with the array of sensing sites; and
normalizing the measured fluorescence emitted by the first detection molecule using the measured fluorescence emitted by the one or more secondary detection molecules.
2. The method of claim 1 , wherein the step of normalizing the measured fluorescence emitted by the first detection molecule comprises dividing the measured fluorescence emitted by the first detection molecule by the measured fluorescence emitted by the one or more secondary detection molecules.
3. The method of claim 1 , wherein the first detection molecule, the one or more secondary detection molecules, the first capture molecule, and the one or more secondary capture molecules are antibodies.
4. The method of claim 1 , wherein the step of normalizing the measured fluorescence emitted by the first detection molecule further comprises generating one or more calibration curves from the measured fluorescence emitted by the one or more secondary detection molecules.
5. The method of claim 1 , wherein the one or more normalizing moieties comprises at least a first normalizing moiety and a second normalizing moiety, wherein the amount of the first normalizing moiety added to the sample is greater than the amount of the second normalizing moiety added to the sample.
6. The method of claim 1 , wherein the step of normalizing the measured fluorescence emitted by the first detection molecule comprises dividing the measured fluorescence emitted by the first detection molecule by a slope of the measured fluorescence emitted by the one or more secondary detection molecules.
7. The method of claim 1 , wherein the step of normalizing the measured fluorescence emitted by the first detection molecule comprises dividing a slope of the measured fluorescence emitted by the first detection molecule by a slope of the measured fluorescence emitted by the one or more secondary detection molecules.