A novel approach for the detection of DNA using immobilized peptide nucleic acid (PNA) probes and signal enhancement by real-time immuno-polymerase chain reaction (RT-iPCR).
A new approach for the detection of DNA target molecules is described, using capture probes and subsequent signal enhancement by a uniform polymerase chain reaction (PCR). Peptide nucleic acid probes were immobilized in real-time PCR-compatible microtiter plates. After hybridization of biotinylated DNA targets, detection was performed by real-time immuno-PCR, a method formerly used for protein detection. We demonstrate the feasibility of this strategy for the qualitative detection of DNA oligonucleotides with a detection limit (LOD) of 6 attomol. Furthermore, the method was applied to PCR-amplified samples from genetically modified maize DNA (Mon810). A 483-bp DNA fragment was detected in mixture with 99.9% of noncomplementary DNA with a sensitivity down to the level of attomole.
Roth
Lillian
L
Bundesinstitut für Risikobewertung (BfR), Abteilung Lebensmittelsicherheit, Thielallee 88-92, 14195, Berlin, Germany. lillian.roth@gmail.com
Zagon
Jutta
J
Ehlers
Anke
A
Kroh
Lothar W
LW
Broll
Hermann
H
D004247:DNA / Q000032:analysis* / Q000276:immunology*; D017930:Genes, Reporter / Q000235:genetics; D015341:Nucleic Acid Probes / Q000032:analysis*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D015341:Nucleic Acid Probes; D004247:DNA