OLFR90 specificity and methods of detection
The present invention describes methods of using Olfr90 demonstrated to bind to fungal metabolites, including a metabolite known to be detected in patients with mold (e.g. Aspergillus ) infections.
1. A detection method comprising the following steps:
a) providing a sample taken from a source to be tested for the presence of a Olfr90 ligand;
b) providing an Olfr90 or a ligand binding part thereof;
c) placing the sample in contact with Olfr90 or the ligand binding part thereof; and
d) detecting the presence or absence of an Olfr90 ligand in the sample, wherein the ligand binding part of Olfr90 binds to 1-Octen-3-ol; 2-Methyl-4-propyl-1,3-oxatiane; 2-Pentylfuran; Allylbenzene; Cinnamaldehyde; 2-Octanone; 3-Octanol; Benzyl Cyanide; Linalool or a combination thereof.
2. The method of claim 1 , wherein the source may be air, water, food, soil, wall scrapings, blood, urine respiratory samples.
3. The method of claim 1 wherein the ligand binding part of Olfr90 is provided.
4. The method of claim 1 , wherein the Olfr90 or the ligand binding part thereof is attached to a reporter and the detecting the present or absence of an Olfr90 ligand in the sample occurs by the displacement of the reporter if the Olfr90 ligand is present in the sample.
5. The method of claim 1 wherein Olfr90 or the ligand binding part thereof is expressed in eukaryotic cells along with a firefly luciferase whose activity is dependent upon elevated [c-AMP] whereby the detecting the presence or absence of an Olfr90 ligand occurs by binding of the Olfr90 ligand to the Olfr90 or ligand binding part thereof elevates the concentration of [cAMP] leading to increased transcription of firefly luciferase if an Olfr90 ligand is present in the sample.
6. The method of claim 1 wherein Olfr90 or the ligand binding part thereof is expressed in eukaryotic cells and a CNG channel (CNGA2) is expressed exogenously, whereby the detecting the presence or absence of an Olfr90 ligand occurs by binding of the Olfr90 ligand to the Olfr90 or ligand binding part thereof elevates the concentration of [cAMP] leading to increased cAMP binding to CNGA2 and a resulting calcium influx may be measured via a detector if an Olfr90 ligand is present in the sample.
7. The method of claim 6 wherein the detector is a calcium sensitive dye or a FLIPR assay.
8. The method of claim 1 wherein in step b) the ligand binding part of Olfr90 is provided and immobilized to a surface.
9. The method of claim 8 further comprising the steps:
incubate the sample with the ligand binding part of Olfr90 to form a mixture; and
add a Olfr90 ligand with a label.
10. The method of claim 9 wherein the label is selected from the group consisting of a fluorescent tag, colorimetric tag, or absorbance tag.
11. The method of claim 1 wherein in step b the ligand binding part of Olfr90 is an aptamer representing the binding site of Olfr90 and the aptamer is labeled with a tag.
12. The method of claim 11 wherein the tag is a fluorophore.
13. The method of claim 12 wherein the aptamer is attached to a surface.
14. The method of claim 13 wherein detecting the presence or absence of an Olfr90 ligand in the sample is determined by the amount of fluorophore that remains bound to the aptamer.
15. The method of claim 14 wherein the surface is a strip.
16. The method of claim 15 wherein the strip is a dip stick.
17. The method of claim 1 wherein the Olfr90 ligand is selected from the group 2-Methyl-4-Propyl-1,3-Oxathiane (MPOT), Cinnamaldehyde, Allybenzene, or combinations thereof.