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- Parkkinen, S, et al.
(författare)
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Sandwich hybridization in solution : a rapid method to screen HPV 16 DNA in cervical scrapes
- 1989
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Ingår i: Molecular and Cellular Probes. - : Elsevier BV. - 0890-8508 .- 1096-1194. ; 3:1, s. 1-11
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Tidskriftsartikel (refereegranskat)abstract
- A solution hybridization method is introduced as a rapid diagnostic method for demonstration of papillomavirus DNA in cervical scrapes. 32P-Labelled detector probe and the biotinylated capture probes were hybridized with DNA of the specimen after pretreatment by boiling in alkaline SDS. After 4 h of hybridization the hybrids were collected onto avidin coated beads and measured. The sensitivity of the method was 1–5 × 105 HPV 16 DNA molecules. Cervical carcinoma cell lines CaSki and SiHa were informative as to the sensitivity of the solution hybridization and the in situ hybridization methods. CaSki cells containing about 700 HPV 16 DNA copies per cell were positive by both methods. SiHa cells with one HPV 16 DNA copy per cell were positive by the sandwich assay but remained negative in the in situ test. A series of 126 cervical scrapes collected from consecutive patients participating in a follow-up study for cervical HPV infection were tested for HPV 16 DNA by both methods. The detection rate of the sandwich test was 19/126 (15%) and that of the in situ method 21/126 (17%) yielding 26 diagnoses altogether. Twelve of these were obtained by one method only. The results obtained by studying the cervical cell lines and repeated specimens taken from constantly HPV 16 positive patients suggested that the two methods can measure different types of infections and thus complement each other in the diagnosis of cervical HPV infections.
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| 2. |
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| 3. |
- Syvänen, Ann-Christine, et al.
(författare)
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Time-resolved fluorometry : a sensitive method to quantify DNA-hybrids
- 1986
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Ingår i: Nucleic Acids Research. - 0305-1048 .- 1362-4962. ; 14:2, s. 1017-1028
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Tidskriftsartikel (refereegranskat)abstract
- Europium and other lanthanides can be excitated with UV-radiation, whereafter the energy is released as fluorescence, delayed in time up to 1 ms after the excitation. Eu can be used as a sensitive label in biological assays. Here we report on the application of time-resolved fluorometry to detect nucleic acid hybrids. The probe DNA was tagged with a hapten, either a fluorene or a sulfone group. After hybridization the probe DNA was detected by a two-step immunological assay with the second antibody labelled with Eu. The method is quantitative with a detection limit of 0.3 pg of actual target regions of immobilized adenovirus genomic DNA. The label was also used in sandwich hybridization, which allowed analyzing nasopharyngeal mucus for the presence of adenovirus.
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