Non-invasive in vivo optical imaging method

A non-invasive method of quantifying the blood level of a fluorescent analyte which comprises a fluorescent entity and a second entity, in a subject, comprising the steps of:

• (a) directing excitation light of at least one predetermined wavelength onto a delineated region comprising at least a portion of the pupil of said subject, to excite the fluorescent entity,
• (b) receiving light emitted from said fluorescent analyte with a wavelength distinguishable from the predetermined wavelength of (a), through the eye of said subject, thereby quantifying the blood level of a fluorescent analyte.

A non-invasive method of determining the presence of a fluorescent analyte which comprises a fluorescent entity and a second entity, in the blood of a subject, comprising or consisting of the steps:

• (a) directing excitation light of at least one predetermined wavelength onto a delineated region comprising at least a portion of the pupil of said subject, to excite the fluorescent entity,
• (b) receiving light emitted from said fluorescent analyte with a wavelength distinguishable from the predetermined wavelength of (a), through the eye of said subject, thereby determining the presence of said fluorescent analyte in the blood of said subject.

The method of any one of the preceding claims, wherein said second entity comprises at least one epitope binding domain which is specific for a target.

The method of any one of the preceding claims, wherein said light which is emitted from said fluorescent analyte with a wavelength distinguishable from the predetermined wavelength of (a), is exclusively received through the eye of said subject.

The method of any one of the preceding claims, wherein in (a) said excitation light of at least one predetermined wavelength is exclusively directed onto a delineated region comprising at least a portion of the pupil of said subject.

The method of any one of the preceding claims, further comprising step (c) determining light emitted from said fluorescent analyte with a wavelength distinguishable from the predetermined wavelength of (a), from further regions of the subject.

The method of any one of the preceding claims, wherein the optical axis of the excitation light of at least one predetermined wavelength which is directed onto a delineated region comprising at least a portion of the pupil of the subject to excite the fluorescent analyte, and the optical axis of the light emitted from said fluorescent analyte with a wavelength distinguishable from the predetermined wavelength, are arranged at an angle to each other.

The method of any one of the preceding claims, wherein the excitation means which is used to direct the excitation light of at least one predetermined wavelength onto a delineated region comprising at least a portion of the pupil of the subject to excite the fluorescent analyte, and the optical detector which is used to receive the light emitted from said fluorescent analyte with a wavelength distinguishable from the predetermined wavelength, are spatially separated.

The method of any one of the preceding claims which is for determining the presence, quantifying the blood level, monitoring or determining the blood clearance of a drug, an antibody or a functional fragment thereof, a protein, a peptide, an enzyme, a nucleic acid for example siRNA, a polysaccharide, a lipid, a receptor, a receptor ligand, a pathogen, a virus, a bacterium, a cell, a cellular target, and/or a tumor antigen in the blood of a subject, for determining the dissolution kinetic of a pharmaceutical or diagnostic composition, and/or for evaluating the elimination pathway of a substance.

The method of any one of the preceding claims, wherein in (b) said light with a wavelength distinguishable from the predetermined wavelength of (a) is received with an optical detector.