Methods and/or use of oligonucleotide conjugates for assays and flow cytometry detections

A live cell flow cytometry method for detecting a target of a sample comprising living cells, comprising:

• i) mixing the sample comprising living cells with a molecular probe and a detectable component, wherein:
• 1) the molecular probe comprises a binding moiety conjugated to a first oligonucleotide sequence, wherein
• a. the binding moiety comprises an antibody, an antibody-fragment, an enzyme, a protein, a peptide, a carbohydrate, a nuclear receptor, a chelator, or combinations or derivatives thereof;
• and
• b. the conjugation between the binding moiety and the first oligonucleotides comprises one or more covalent bond linkages selected from the group consisting of a hydrazine, an oxime, and a triazine;
• and
• 2) the detectable component is not immobilized on a solid support and comprises a signal generator conjugated to a second oligonucleotide having a sequence complementary to the sequence of the first oligonucleotide, wherein
• a. the signal generator comprises a scaffold molecule further conjugated to more than one signal generating moieties, the scaffold molecule comprising a dendrimer, a polysaccharide, or combinations or derivatives thereof;
• and
• b. the second oligonucleotide is directly conjugated to the scaffold molecule of the signal generator via one or more covalent bond linkages selected from the group consisting of a hydrazine, an oxime, and a triazine;
• ii) binding the target in the sample with the binding moiety of the molecular probe;
• iii) hybridizing the first oligonucleotide with the second oligonucleotide;
• iv) reducing the potential for cross-talk by adding to the molecular probe and the detectable component an oligonucleotide comprising at least one of:
• 1) an unconjugated oligonucleotide having a sequence complementary to the sequence of the first oligonucleotide;
• and
• 2) an unconjugated oligonucleotide having a sequence complementary to the sequence of the second oligonucleotide;
• then
• v) detecting a signal generated from the detectable component hybridized to the target-bound molecular probe using flow cytometry, wherein the hybridized detectable component is not immobilized on a solid support;
• and
• vi) removing the hybridized detectable component from the bound target, wherein the removal is by a stripping and washing process.