Nucleic acid multimers and amplified nucleic acid hybridization assays using same
DCFirst Claim
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1. A synthetic branched nucleic acid multimer having a minimum of three termini comprising a combination of 5'"'"' and 3'"'"' termini comprising:
- (a) at least one first single-stranded oligonucleotide unit that is capable of hybridizing specifically to a first single-stranded nucleic acid sequence of interest; and
(b) a multiplicity of second single-stranded oligonucleotide units that are capable of hybridizing specifically to a second single-stranded nucleic acid sequence of interest, wherein the first single-stranded oligonucleotide unit is bonded directly or indirectly to the multiplicity of second single-stranded oligonucleotide units only via covalent bonds.
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Abstract
Linear or branched oligonucleotide multimers useful as amplifiers in biochemical assays which comprise (1) at least one first single-stranded oligonucleotide unit that is complementary to a single-stranded oligonucleotide sequence of interest, and (2) a multiplicity of second single-stranded oligonucleotide units that are complementary to a single-stranded labeled oligonucleotide. Amplified sandwich nucleic acid hybridizations and immunoassays using the multimers are exemplified.
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Citations
59 Claims
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1. A synthetic branched nucleic acid multimer having a minimum of three termini comprising a combination of 5'"'"' and 3'"'"' termini comprising:
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(a) at least one first single-stranded oligonucleotide unit that is capable of hybridizing specifically to a first single-stranded nucleic acid sequence of interest; and (b) a multiplicity of second single-stranded oligonucleotide units that are capable of hybridizing specifically to a second single-stranded nucleic acid sequence of interest, wherein the first single-stranded oligonucleotide unit is bonded directly or indirectly to the multiplicity of second single-stranded oligonucleotide units only via covalent bonds. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 38, 57)
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37. A solution sandwich nucleic acid hybridization assay for detecting a first nucleic acid sequence that is part of a nucleic acid segment that includes a second nucleic acid sequence in a sample that contains said segment and another nucleic acid segment that comprises the first nucleic acid sequence but does not include the second nucleic acid sequence comprising:
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(a) contacting the analyte under hybridizing conditions with an excess of (i) an amplifier probe oligonucleotide comprising a first segment that is complementary to one of the first nucleic acid sequence or the second nucleic acid sequence and a second segment that is complementary to an oligonucleotide unit of a nucleic acid multimer and (ii) a capture probe oligonucleotide comprising a first segment that is complementary to the other of the first nucleic acid sequence or the second nucleic acid sequence and a second segment that is complementary to an oligonucleotide bound to a solid phase; (b) contacting under hybridizing conditions the product of step (a) with the oligonucleotide bound to the solid phase; (c) thereafter separating materials not bound to the solid phase; (d) contacting under hybridizing conditions the solid phase complex product of step (c) with the nucleic acid multimer, said multimer comprising (i) at least one oligonucleotide unit that is complementary to the second segment of the amplifier probe oligonucleotide and (ii) a multiplicity of second oligonucleotide units that are complementary to a labeled oligonucleotide; (e) removing unbound multimer; (f) contacting under hybridizing conditions the solid phase complex product of step (e) with the labeled oligonucleotide; (g) removing unbound labeled oligonucleotide; and (h) detecting the presence of label in the solid phase complex product of step (g).
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39. A synthetic linear nonhomopolymeric nucleic acid multimer useful as a means for amplifying a detectable signal in an assay involving nucleic acid hybridization consisting essentially of:
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(a) at least one first single-stranded oligonucleotide unit that is capable of hybridizing specifically to a first single-stranded nucleic acid sequence of interest; and (b) a multiplicity of second single-stranded oligonucleotide units all of which are capable of hybridizing specifically to a second single-stranded nucleic acid sequence of interest, wherein the first single-stranded oligonucleotide unit is bonded directly or indirectly to the multiplicity of second single-stranded oligonucleotide units only via covalent bonds. - View Dependent Claims (40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 58, 59)
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Specification