Cell-free biosensors to detect creatinine, creatine, and sarcosine
AssigneeGeorgia Tech Research Corporation
InventorsMark Styczynski, Elizabeth Hutson Chilton, Emily Heckard
Life of this patent
Estimating the term…
Abstract
An exemplary embodiment of the present disclosure provides a biosensor that can facilitate detection of the molecules creatinine, creatine and/or sarcosine in a minimal-equipment, portable, and low-cost fashion. Some aspects of the disclosure relate to methods and systems to enable quantitative measurement of certain biomarkers at the point of care without expensive instrumentation.
Claims17
A method of generating a diagnostic tool for measuring an unknown amount of an analyte in a biological sample using a cell-free extract (CFE), the diagnostic tool comprising a plurality of reference points, each reference point being a distinct color and corresponding to a different amount of the analyte, the method comprising:
- determining a desired amount of a regulator of a reporter, the regulator comprising a sarcosine-responsive transcription factor from Pseudomonas aeruginosa (SouR);
- determining a saturating amount of the analyte;
- and
- determining a desired reaction time;
- wherein the desired amount of the regulator of the reporter, the saturating amount of the analyte, and the desired reaction time are determined such that when the unknown amount of the analyte is combined with the biological sample, the CFE, and the desired amount of the regulator of the reporter for the desired reaction time, a colorimetric gene product is generated having a color corresponding to a color of a first reference point in the plurality of reference points containing the biological sample, the CFE, the saturating amount of analyte, and the desired amount of the regulator of the reporter, the first reference point associated with an amount of the analyte about equal to the unknown amount of analyte.
A method for generating a range of visible colors in serum samples with unknown amounts of an analyte by addition of a small molecule, wherein the small molecule displaces an interfering molecule from one or more of a regulator, a reporter, and a molecule acted on by the reporter, the method comprising:
- providing a desired amount of a regulator of a reporter, the regulator comprising a sarcosine-responsive transcription factor from Pseudomonas aeruginosa (SouR);
- providing a saturating amount of the analyte;
- and
- providing a desired reaction time;
- wherein:
- the desired amount of the regulator of the reporter, the saturating amount of the analyte, and the desired reaction time are determined such that when the unknown amount of the analyte is combined with the serum and the desired amount of the regulator of the reporter for the desired reaction time, a colorimetric gene product is generated having a color corresponding to a color of a first reference point in a plurality of reference points containing the serum, the saturating amount of analyte, and the desired amount of the regulator of the reporter, the first reference point associated with an amount of the analyte about equal to the unknown amount of analyte;
- and
- the method is configured to reduce or eliminate inter-sample variability.
A method for generating a range of visible colors in serum samples with unknown amounts of an analyte by addition of a small molecule, wherein the small molecule displaces an interfering molecule from one or more of a regulator, a reporter, and a molecule acted on by the reporter, the method comprising:
- providing a desired amount of a reporter, the reporter comprising one or more plasmids;
- providing a saturating amount of the analyte;
- providing a desired reaction time;
- and
- stabilizing the reporter, the stabilizing comprising providing a stabilizing compound selected from the group consisting of doxorubicin, vitamin C, citric acid, vitamin E, uric acid, L-glutathione (reduced), and (±)-6-Hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid and combinations thereof;
- wherein:
- the desired amount of the reporter, the saturating amount of the analyte, and the desired reaction time are determined such that when the unknown amount of the analyte is combined with the serum and the desired amount of the reporter for the desired reaction time, the reporter generates a color corresponding to a color of a first reference point in a plurality of reference points containing the serum, the saturating amount of analyte, and the desired amount of the reporter, the first reference point associated with an amount of the analyte about equal to the unknown amount of analyte;
- and
- the method is configured to reduce or eliminate inter-sample variability.
Dependencies are inferred from the claim text.