3D-exoquant method for the analysis of surface molecules and quantification of tissue-specific exosomes in biological fluids

A method of identifying and/or quantifying one or more antigens of interest (biomarkers) on the surface of cell- or tissue-specific exosomes, said method comprising:

• i) incubating a population of exosomes obtained from a biological fluid with a one or more tissue-specific antibodies that bind an antigen specific to a tissue or cell type of interest that produces exosomes, where said tissue specific antibodies are attached to acceptor beads and said incubating is under conditions where said tissue-specific antibodies bind exosomes displaying said antigen;
• ii) isolating exosomes bound by said tissue specific antibodies to produce isolated tissue-specific exosomes;
• iii) incubating a test subset of said isolated tissue-specific exosomes with acceptor beads attached to test antibodies that bind an antigen of interest under conditions where said antibodies that bind an antigen of interest bind to exosomes that display said antigen of interest;
• iv) incubating a control subset of said isolated tissue-specific exosomes with negative control acceptor beads under the same conditions as in step iii where the concentration of control acceptor beads is about the same as the concentration of the acceptor beads attached to an antibody that binds a marker of interest;
• v) incubating said test subset of isolated exosomes and said control subset of said exosomes with an antibody that binds an exosome specific antigen where said antibody is attached to a donor that produces singlet oxygen when illuminated;
• vi) detecting a signal produced upon illumination of said control subset and said test subset where said signal is produced by reaction of the negative control acceptor beads reaction of the acceptor beads attached to test antibodies with said donor;
• and
• vii) identifying the presence or quantifying said antigen(s) of interest, wherein said identifying or quantifying comprises determining the difference in signal produced upon illumination of said test subset and said control subset.