Method and marker for analyzing a sample

38:

• A linker, configured to bind to an affinity reagent, the linker comprising:
• at least one binding site configured to bind at least one dye;
• and
• an oligonucleotide attachment portion configured to bind to an oligonucleotide barcode of the affinity reagent.

40:

• A reporter, comprising:
• a linker according to claim 38;
• and
• the at least one dye bound to the at least one binding site of the linker.

41:

• A reporter, comprising:
• a linker according to claim 39;
• and
• a combination of dyes, each respective dye bound to a respective one of the plurality of binding sites of the linker, wherein at least two of the combination of dyes have different characteristics for at least one of:
• excitation or emission.

42:

• A marker, comprising:
• a reporter according to claim 40;
• and
• the affinity reagent attach to an analyte, wherein the reporter is attached to the affinity reagent.

52:

• A method for analyzing a sample, the sample comprising:
• a plurality of affinity reagents, each respective affinity reagent being configured to attach to a respective analyte, at least one of the plurality of affinity reagents being attached to an analyte;
• and
• a first plurality of combinations of dyes, each respective combination of dyes being unique among the first plurality of combinations of dyes, and each respective combination of dyes comprising at least two dyes having different characteristics for at least one of excitation or emission,
• wherein at least one of the first plurality of combinations of dyes is attached to an associated affinity reagent according to a first mapping,
• wherein at least another one of the first plurality of combinations of dyes is attached to the associated affinity reagent according to a second mapping,
• wherein the first mapping is different from the second mapping,
• the method comprising, for the first mapping and for the second mapping in a time sequential manner:
• i) directing excitation light at the sample, the excitation light having characteristics for exciting the at least two dyes having different characteristics for at least one of:
• excitation or emission;
• and
• ii) generating at least one first readout from emission light emitted by the at least two dyes excited by excitation light.

53:

• A method for analyzing a sample, the sample comprising:
• a plurality of affinity reagents, each respective affinity reagent being configured to attach to a respective analyte, at least one of the plurality of affinity reagents being attached to an analyte;
• and
• a first plurality of combinations of dyes, each combination of dyes of the first plurality of combinations of dyes being unique among the first plurality of combinations of dyes and comprising at least two dyes having different characteristics for at least one of:
• excitation or emission,
• a second plurality of combinations of dyes, each combination of dyes of the second plurality of combination of dyes being unique among the second plurality of combinations of dyes and comprising at least two dyes having different characteristics for at least one of excitation or emission,
• wherein the second plurality of combinations of dyes is different from the first plurality of combinations of dyes,
• wherein at least one of the first plurality of combinations of dyes is attached to an associated affinity reagent according to a first mapping,
• wherein at least one of the second plurality of combinations of dyes is attached to the associated affinity reagent according to a second mapping,
• wherein the first mapping is different from the second mapping,
• the method comprising, for the first mapping and for the second mapping in a time sequential manner:
• i) directing excitation light at the sample, the excitation light having characteristics for exciting the at least two dyes having different characteristics for at least one of:
• excitation or emission;
• and
• ii) generating at least one first readout from emission light emitted by the at least two dyes excited by the excitation light.

55:

• The method according to claim 52, further comprising at least one of deactivating at least one dye in the first plurality of combinations of dyes;
• removing an attachment between at least one affinity reagent and at least one of the first plurality of combinations of dyes;
• removing an attachment between at least one affinity reagent and at least one of the analytes;
• or
• waiting longer than a fluorescence lifetime of at least one dye in the first plurality of combinations of dyes, and repeating steps i) and ii) for the first plurality of combinations of dyes according to the second mapping.

56:

• The method according to claim 52, wherein the at least another one of the first plurality of combinations of dyes is stochastically attached to the associated affinity reagent according to the second mapping, and wherein stochastic labeling is achieved by at least one of DNA-PAINT, DNA-Exchange, Exchange-PAINT, or by using oligonucleotide barcodes attached to the affinity reagents and complementary attachment oligonucleotide portions attached to the first plurality of combinations of dyes, which are configured to allow repeated association and dissociation of the first plurality of combinations of dyes and the affinity reagents by at least one of the following:
• a change of concentration of a host molecule;
• a change of temperature;
• a change of illumination condition;
• a change in a concentration of at least one capture strand;
• or
• a change in a concentration and/or activity of an enzyme.

57:

• The method according to claim 52, further comprising:
• generating a second readout from the emission light;
• and
• determining, by at least one computer processor, at least a first affinity reagent present in the sample, by:
• comparing the at least one first readout and the second readout;
• and
• determining a presence of the first affinity reagent based, at least in part, on the comparison.