Use of Up-Converting Phosphor Reporters in Lateral-Flow Assays to Detect Specific Nucleic Acid Sequences: A Rapid, Sensitive DNA Test to Identify Human Papillomavirus Type 16 Infection

• Published in: Clinical chemistry (Baltimore, Md.) Md.), 2001-10, Vol.47 (10), p.1885-1893
• Main Authors: Corstjens, Paul, Zuiderwijk, Michel, Brink, Antoinette, Li, Shang, Feindt, Hans, Niedbala, R. Sam, Tanke, Hans
• Format: Article
• Language: English
• Subjects: Antibodies, Monoclonal; Base Sequence; Biological and medical sciences; DNA, Viral - genetics; DNA, Viral - immunology; Erbium; Female; Fluorescence; Haptens; Humans; Investigative techniques, diagnostic techniques (general aspects); Medical sciences; Miscellaneous. Technology; Nucleic Acid Hybridization - methods; Papillomaviridae - genetics; Papillomavirus Infections - diagnosis; Pathology. Cytology. Biochemistry. Spectrometry. Miscellaneous investigative techniques; Polymerase Chain Reaction - instrumentation; Polymerase Chain Reaction - methods; Sensitivity and Specificity; Tumor Virus Infections - diagnosis; Uterine Cervical Neoplasms - virology; Ytterbium
• ISSN: 0009-9147; 1530-8561
• DOI: 10.1093/clinchem/47.10.1885

A lateral-flow (LF) device using the new reporter up-converting phosphor technology (UPT) was applied to DNA (hybridization) assays for the detection of specific nucleic acid sequences, thereby aiming to perform the test outside well-equipped laboratories. The methodology reported here is sensitive and provides a rapid alternative for more elaborate gel electrophoresis and Southern blotting. In a preliminary study, it was applied to screen for the presence of human papillomavirus type 16 (HPV16) in a defined series of cervical carcinomas. A LF assay was used to capture haptenized DNA molecules and hybrids, which were immunolabeled (before LF) with 400-nm UPT particles. These particles emit visible light after excitation with infrared in a process called up-conversion. Because up-conversion occurs in only the phosphor lattice, autofluorescence of other assay components is virtually nonexistent. The use of the UPT reporter in LF-DNA tests, as compared with colloidal gold, improved the detection limit at least 100-fold. UPT LF-DNA tests were successfully applied to detect (in a blind test) the presence of HPV16 in DNA extracts obtained from cervical carcinomas. Test results matched 100% with previous characterization of these carcinomas. The use of UPT in LF assays to detect specific nucleic acids provides low attamole-range sensitivity. Hybridization and consecutive detection of PCR-amplified HPV16 sequences were successful in a background of 10 microg of fish-sperm DNA. The sensitivity of UPT detection in these complex mixtures indicates that detection of viral infections without PCR or other amplification technique is achievable.