Amplification-free detection of Cryptosporidium parvum nucleic acids with the use of DNA/RNA-directed gold nanoparticle assemblies

S. E. Weigum, Alejandro Castellanos, A. Clinton White, R. Richards-Kortum

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

This study describes the development and evaluation of an amplification-free molecular assay for detection of Cryptosporidium parvum oocysts. The assay employed a pair of oligonucleotide-functionalized gold nanoparticle (AuNP) probes that were complementary to adjacent sequences on C. parvum 18s rRNA. Hybridization of the probes to the target RNA resulted in the assembly of AuNPs into target-linked networks, which were detected both visibly and spectroscopically, by a redshift in the wavelength of light scattered by the gold nanoparticles. The limit of detection was between 4 × 105 and 4 × 106 copies of RNA per microliter reaction mix, when a short synthetic target or full-length in vitro transcribed target was employed. With total nucleic acids purified from C. parvum oocysts spiked into 100-mg stool, as few as 670 oocysts/μl reaction mix were detected. The ability to detect the nucleic acids of C. parvum oocysts in stool, without the need for complex amplification, offers unique advantages for such AuNP aggregation assays to be extended toward use in resource-limited settings where protozoan detection is needed most.

Original languageEnglish (US)
Pages (from-to)923-926
Number of pages4
JournalJournal of Parasitology
Volume99
Issue number5
DOIs
StatePublished - 2013

Fingerprint

Cryptosporidium parvum
Oocysts
nucleic acid
oocysts
Gold
Nanoparticles
Nucleic Acids
nucleic acids
RNA
amplification
gold
assay
DNA
probe
assays
oligonucleotides
Oligonucleotides
wavelength
Protozoa
Limit of Detection

ASJC Scopus subject areas

  • Parasitology
  • Ecology, Evolution, Behavior and Systematics

Cite this

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title = "Amplification-free detection of Cryptosporidium parvum nucleic acids with the use of DNA/RNA-directed gold nanoparticle assemblies",
abstract = "This study describes the development and evaluation of an amplification-free molecular assay for detection of Cryptosporidium parvum oocysts. The assay employed a pair of oligonucleotide-functionalized gold nanoparticle (AuNP) probes that were complementary to adjacent sequences on C. parvum 18s rRNA. Hybridization of the probes to the target RNA resulted in the assembly of AuNPs into target-linked networks, which were detected both visibly and spectroscopically, by a redshift in the wavelength of light scattered by the gold nanoparticles. The limit of detection was between 4 × 105 and 4 × 106 copies of RNA per microliter reaction mix, when a short synthetic target or full-length in vitro transcribed target was employed. With total nucleic acids purified from C. parvum oocysts spiked into 100-mg stool, as few as 670 oocysts/μl reaction mix were detected. The ability to detect the nucleic acids of C. parvum oocysts in stool, without the need for complex amplification, offers unique advantages for such AuNP aggregation assays to be extended toward use in resource-limited settings where protozoan detection is needed most.",
author = "Weigum, {S. E.} and Alejandro Castellanos and White, {A. Clinton} and R. Richards-Kortum",
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T1 - Amplification-free detection of Cryptosporidium parvum nucleic acids with the use of DNA/RNA-directed gold nanoparticle assemblies

AU - Weigum, S. E.

AU - Castellanos, Alejandro

AU - White, A. Clinton

AU - Richards-Kortum, R.

PY - 2013

Y1 - 2013

N2 - This study describes the development and evaluation of an amplification-free molecular assay for detection of Cryptosporidium parvum oocysts. The assay employed a pair of oligonucleotide-functionalized gold nanoparticle (AuNP) probes that were complementary to adjacent sequences on C. parvum 18s rRNA. Hybridization of the probes to the target RNA resulted in the assembly of AuNPs into target-linked networks, which were detected both visibly and spectroscopically, by a redshift in the wavelength of light scattered by the gold nanoparticles. The limit of detection was between 4 × 105 and 4 × 106 copies of RNA per microliter reaction mix, when a short synthetic target or full-length in vitro transcribed target was employed. With total nucleic acids purified from C. parvum oocysts spiked into 100-mg stool, as few as 670 oocysts/μl reaction mix were detected. The ability to detect the nucleic acids of C. parvum oocysts in stool, without the need for complex amplification, offers unique advantages for such AuNP aggregation assays to be extended toward use in resource-limited settings where protozoan detection is needed most.

AB - This study describes the development and evaluation of an amplification-free molecular assay for detection of Cryptosporidium parvum oocysts. The assay employed a pair of oligonucleotide-functionalized gold nanoparticle (AuNP) probes that were complementary to adjacent sequences on C. parvum 18s rRNA. Hybridization of the probes to the target RNA resulted in the assembly of AuNPs into target-linked networks, which were detected both visibly and spectroscopically, by a redshift in the wavelength of light scattered by the gold nanoparticles. The limit of detection was between 4 × 105 and 4 × 106 copies of RNA per microliter reaction mix, when a short synthetic target or full-length in vitro transcribed target was employed. With total nucleic acids purified from C. parvum oocysts spiked into 100-mg stool, as few as 670 oocysts/μl reaction mix were detected. The ability to detect the nucleic acids of C. parvum oocysts in stool, without the need for complex amplification, offers unique advantages for such AuNP aggregation assays to be extended toward use in resource-limited settings where protozoan detection is needed most.

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