Methods for detecting multiple nucleic acids in a sample using reporter compounds and binding members thereof

A method of determining the presence and/or amount of nucleic acids in a sample, comprising

• (a) providing
• an amount of a first nucleic acid;
• an amount of a second nucleic acid;
• an amount of a FRET compound capable of forming complexes with the first nucleic acid, wherein the FRET compound comprises a binding portion specific to a region of the first nucleic acid and a first detectable label that has, in an uncomplexed state, a first detectable characteristic and, in a complexed state, a second detectable characteristic that differs from the first detectable characteristic;
• an amount of a reporter compound capable of forming complexes with the second nucleic acid, wherein each reporter compound comprises a binding portion specific to a region of the second nucleic acid and a second detectable label;
• and
• a binding member capable of capturing the reporter compound, wherein forming of complexes of the second nucleic acid with the reporter compound inhibits capturing of the reporter compound by the binding member;
• (b) subjecting the first and/or second nucleic acid to amplification;
• (c) forming complexes each comprising a first nucleic acid and a FRET compound;
• (d) forming complexes of a subset of the amount of reporter compound with a second nucleic acid;
• (e) capturing a remaining subset of the amount of reporter compound not in complex with a second nucleic acid on the binding member;
• (f) determining a value indicative for the presence and/or amount of first nucleic acid captured by FRET compounds;
• and
• (g) determining a value indicative for the presence and/or amount of reporter compound captured on the binding member.