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Detection of plurality of targets in biological samples

  • US 9,677,125 B2
  • Filed: 10/21/2009
  • Issued: 06/13/2017
  • Est. Priority Date: 10/21/2009
  • Status: Active Grant
First Claim
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1. A method of detecting at least three sets of different targets in a biological sample via simultaneous addition of at least three sets of different target-binding probes followed by their sequential detection, the method comprising:

  • (a) simultaneously contacting the at least three sets of different targets with the at least three sets of different target-binding probes to form a plurality of target-bound probes;

    (b) covalently attaching each of the plurality of target-bound probes of step (a) to the corresponding target or to a molecule that is located in the vicinity of the corresponding target in the biological sample to form a plurality of covalently attached probes; and

    (c) sequentially detecting the plurality of covalently attached probes of step (b) to sequentially detect each set of different target, wherein the sequential detection comprises;

    (i) generating a first set of signals from a first set of probes in the plurality of covalently attached probes by contacting a first set of signal-generating probes that are capable of binding to the first set of probes;

    (ii) detecting a first set of targets via observing the first set of signals that is generated in step (i);

    (iii) modifying the observed first set of signals by applying a chemical agent after step (ii);

    (iv) generating a second set of signals after step (iii) from a second set of probes in the plurality of covalently attached probes by contacting a second set of signal-generating probes that are capable of binding to the second set of probes;

    (v) detecting a second set of targets via observing the second set of signals that is generated in step (iv);

    (vi) modifying the observed second set of signals by applying a chemical agent after step (v); and

    (vii) generating a subsequent set of signals from a subsequent set of probes in the plurality of covalently attached probes, detecting a subsequent set of targets via observing the subsequent set of signals, modifying the observed subsequent set of signals by applying a chemical agent, and repeating these steps till all sets of different targets are detected.

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