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US 8,465,922 B2Granted
Methods and systems for monitoring reactions
AssigneePacific Biosciences of California, Inc.
InventorsJohn Eid, Stephen Turner
Life of this patent
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Abstract
Methods and systems for monitoring reactions by observing signals deriving from those reactions, using signal processing that allows differentiation between signals that are otherwise optically overlapping by conventional detection methods. Centroid determination is used to identify signal sources that are presenting confounding overlapping signals due to their physical proximity, and/or to identify discrete signals from different reaction centers.
Claims11
A method of monitoring single molecule reactions at a first and a second single molecule reaction complex, comprising:
- providing a first single molecule reaction complex and a second single molecule reaction complex, each at least partially immobilized on a substrate,
- exposing the first and second single molecule reaction complexes to reaction conditions;
- wherein said first single molecule reaction complex produces a first optically detectable signal indicative of the presence of a single molecule reaction at the first single molecule reaction complex and the second single molecule reaction complex produces a second optically detectable signal indicative of the presence of a single molecule reaction at the second single molecule reaction complex, wherein the first and second optically detectable signals are not completely optically resolvable from each other;
- positionally detecting the first optically detectable signal and the second optically detectable signal;
- identifying a first positional centroid for the first optically detectable signal and a second positional centroid for the second optically detectable signal;
- and
- characterizing the first optically detectable signal as emanating from the first single molecule reaction complex based upon the first positional centroid and characterizing the second optically detectable signal as emanating from the second single molecule reaction complex based upon the second positional centroid, and;
- detecting a plurality of optically detectable signals from the first and second single molecule reaction complexes and assigning each of said plurality of signals as emanating from either the first or second single molecule reaction, complex based upon a comparison of a positional centroid of each of the plurality of optically detectable signals with a positional centroid determined from the first and second single molecule reaction complexes.
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