Mitigation of photodamage in analytical reactions
AssigneePacific Biosciences of California, Inc.
InventorsJohn Eid, Devon Murphy, Geoffrey Otto, Stephen Turner
Life of this patent
Estimating the term…
Abstract
Compositions, devices, systems and methods for reducing and/or preventing photodamage of one or more reactants in illuminated analytical reactions by one or more of incorporating photodamage mitigating agents within the reaction mixture and/or interrogating different observation regions of the reaction mixture for a period that is less than a photodamage threshold period.
Claims25
1-28. (canceled)
A method of performing an illuminated reaction, comprising:
- providing a substrate having a reaction mixture disposed thereon, wherein the reaction mixture comprises a first reactant, a second reactant and a photodamage mitigating agent, wherein the photodamage mitigating agent reduces an amount of photodamage to the first reactant resulting from interaction of the first reactant with the second reactant under excitation illumination that would occur in the absence of the photodamage mitigating agent;
- and
- illuminating the reaction mixture on the substrate, with an excitation illumination.
A method of performing an enzyme reaction, comprising:
- providing an enzyme within a first observation region;
- contacting the enzyme with a fluorescent or fluorogenic substrate for the enzyme;
- and
- directing an excitation radiation at and detecting signals from the first observation region for a period that is less than a photodamage threshold period.
A method of monitoring a base extension reaction, comprising:
- providing a polymerase enzyme within a first observation region;
- contacting the polymerase with at least a first fluorescent or fluorogenic nucleotide analog;
- and
- monitoring a fluorescent signal emitted from the first observation region in response to illumination with excitation radiation for a period that is less than a photodamage threshold period.
(canceled)
A method of performing an enzyme reaction, comprising:
- providing an enzyme within an observation region;
- contacting the enzyme with a fluorescent or fluorogenic substrate for the enzyme under excitation illumination, in the presence of at least a first photodamage mitigating agent.
A method of monitoring a base extension reaction, comprising:
- providing a polymerase enzyme within an observation region;
- contacting the polymerase with at least a first fluorescent or fluorogenic nucleotide analog in the presence of at least a first photodamage mitigating agent;
- and
- monitoring a fluorescent signal emitted from the observation region in response to illumination with excitation radiation.
A method of monitoring a reaction mixture comprising at least a first enzyme and a fluorescent or fluorogenic substrate for the first enzyme, comprising directing an excitation radiation at a first observation region for a first period that is less than a photodamage threshold period.
Dependencies are inferred from the claim text.