HPV genotype profile in a Norwegian cohort with ASC-US and LSIL cytology with three year cumulative risk of high grade cervical neoplasia.

OBJECTIVE: To explore the HPVgenotype profile in Norwegian women with ASC-US/LSIL cytology and the subsequent risk of high-grade cervical neoplasia (CIN 3+). METHODS: In this observational study delayed triage of ASC-US/LSIL of 6058 women were included from 2005 to 2010. High-risk HPV detection with Hybrid Capture 2 (HC2) was used and the HC2+ cases were genotyped with in-house nmPCR. Women were followed-up for histologically confirmed CIN3+ within three years of index HPV test by linkage to the screening databases at the Cancer Registry of Norway. RESULTS: HC2 was positive in 45% (2756/6058) of the women. Within 3years CIN3+ was diagnosed in 26% of women<34year and in 15%≥34year. HC2 was positive at index in 94% of CIN3+ cases and negative in 64 cases including three women with cervical carcinomas. Women<34years with single infections of HPV 16, 35, 58 or 33 or multiple infections including HPV 16, 52, 33 or 31 were associated with highest proportions of CIN 3+. Older women with single infection with HPV 16, 33, 31 or 35 or multiple infections including HPV 16, 33, 31 or 18/39 were more likely to develop CIN 3+. CONCLUSIONS: HPV 16 and HPV 33 at baseline both as single or multiple infections, were associated with the highest risk for CIN3+. Among older women, all 13 high-risk genotypes as single infection were associated with >20% risk of CIN3+. Further studies are necessary to risk stratify the individual genotypes to reduce the number of colposcopies in Norway.

50 years of screening in the Nordic countries: quantifying the effects on cervical cancer incidence.

BACKGROUND: Nordic countries' data offer a unique possibility to evaluate the long-term benefit of cervical cancer screening in a context of increasing risk of human papillomavirus infection. METHODS: Ad hoc-refined age-period-cohort models were applied to the last 50-year incidence data from Denmark, Finland, Norway and Sweden to project expected cervical cancer cases in a no-screening scenario. RESULTS: In the absence of screening, projected incidence rates for 2006-2010 in Nordic countries would have been between 3 and 5 times higher than observed rates. Over 60,000 cases or between 41 and 49% of the expected cases of cervical cancer may have been prevented by the introduction of screening in the late 1960s and early 1970s. CONCLUSIONS: Our study suggests that screening programmes might have prevented a HPV-driven epidemic of cervical cancer in Nordic countries. According to extrapolations from cohort effects, cervical cancer incidence rates in the Nordic countries would have been otherwise comparable to the highest incidence rates currently detected in low-income countries.

Large performance variation does not affect outcome in the Finnish cervical cancer screening programme.

OBJECTIVE: Cytology screening for prevention of cervical cancer can reduce incidence and mortality by more than 80% in settings with good organization and rigorous quality control. Audit studies are essential for reaching and maintaining a high quality of screening. The aim of this study was to evaluate variation in performance indicators by screening laboratory and assess the impact on the effectiveness of screening as indicated by cervical intraepithelial neoplasia grade 3 and above (CIN3+) rates after a negative screen. METHODS: Seven cytology screening laboratories operating during 1990-1999 with a total of 953 610 screening tests performed were included in the study. By linking screening and cancer register files, all cases of CIN3+ diagnosed in the screened population were identified. For 395 CIN3+ cases with a preceding negative screen and 787 controls, a re-evaluation of smears was undertaken to uncover false negative screening tests. Performance parameters and rates of CIN3+ after a negative screen were analysed for interlaboratory heterogeneity. RESULTS: The rates of follow-up recommendations and referrals varied by up to 3.6- (2.8-10.2%) and 4.0-fold (0.03-0.12%), respectively. CIN1, CIN2 and CIN3+ screen detection rates differed by up to 8.5- (0.02-0.17%), 5.4- (0.05-0.25%) and 3.3-fold (0.05-0.18%). False negative rates determined by re-evaluation showed up to 2.1-fold differences (29-62%). Rates of CIN3+ after a negative screen (0.023-0.048%) and as a proportion of total CIN3+ (15-31%) in the screened population were low and did not vary significantly. CONCLUSIONS: There were large variations in the sensitivity-specificity trade-off between laboratories, reflected in all performance indicators as well as in the test validity estimates of the re-evaluation phase, but not in screening effectiveness. Even though performance variations do not always have an impact on the effectiveness of screening, they lead to variations in cost, treatment and psychological burden, and should be addressed.

A group-specific microbiological test for the detection of tetracycline residues in raw milk.

The potentiality of using a luminescent Escherichia coli strain for the specific detection of tetracycline residues in raw bovine milk was investigated. The sensor cells contain a reporter plasmid carrying the bacterial luciferase operon of Photorhabdus luminescens under the control of the tetracycline responsive control region from transposon Tn10. Incubation of the cells with the sample containing tetracyclines increases the light emission of the sensor cells. The most sensitive tetracycline detection was achieved in 120 min and by using CDTA as a chelating agent in the assay. Heat-treatment of milk before the assay decreased the variations in background luminescence signals and in tetracycline-induced luminescence between different milk samples. The detection limits for tetracycline, oxytetracycline, chlortetracycline, doxycycline, methacycline, demeclocycline, and minocycline were between 2 and 35 ng/mL. Nontetracycline antibiotics did not significantly interfere with the detection of tetracyclines.

Qualitative detection of tetracycline residues in milk with a luminescence-based microbial method: the effect of milk composition and assay performance in relation to an immunoassay and a microbial inhibition assay.

Performance of Tet-Lux, a newly developed microbiological test for the detection of tetracycline residues in raw milk, based on tetracycline-controlled luminescence activation of the test bacteria, was evaluated in bovine milks with variable amounts of somatic cells, bacteria, fat, protein, and natural inhibitory compounds. The sensitivity of Tet-Lux was also compared to a commercially available tetracycline immunoassay (Snap, Idexx Laboratories Inc.) and to a microbial inhibition test (Delvotest SP, Gist-Brogades). There were slight differences in the luminescence signals between different milk samples, but no single factor could be pointed out to be responsible for them. There appeared to be a modest inverse relationship between luminescence and increasing fat and protein content. The amount of somatic cells, bacteria, and the natural inhibitors lysozyme and lactoferrin did not affect the luminescence response. The test fulfilled the sensitivity requirement specified by the European Union (maximum residue limit 100 ng/ml for tetracyclines). The Tet-Lux test was clearly more sensitive to all tetracyclines tested (oxytetracycline, tetracycline, chlortetracycline, doxycycline, demeclocycline, methacycline, minocycline) than Delvotest SP, and for five tetracyclines out of seven more sensitive than Snap. The test provides a fast, simple, and robust microbial method for the qualitative detection of tetracycline residues in milk.

Zeptomole detection sensitivity of prostate-specific antigen in a rapid microtitre plate assay using time-resolved fluorescence.

Prostate-specific antigen (PSA) was detected in microtitre wells coated with a PSA-specific antibody using biotinylated antibody and streptavidin-coated, highly fluorescent 107 nm nanoparticles, which contained more than 30000 europium ions entrapped by beta-diketones. PSA was monitored directly on the surface of a well without any additional enhancement step. The sensitivity of the assay was 1.6 ng/L, corresponding to 50 fmol/L or 250 zeptomoles (250 x 10(-21) mol/L) of PSA. The high specific activity and low non-specific binding of the streptavidin-coated nanoparticles improved the sensitivity of the PSA assay 100-fold compared to the conventional europium-labelled streptavidin tracer in the same assay format. Additionally, the streptavidin-coated nanoparticle label made very rapid assays possible, due to the high affinity of the streptavidin-biotin complex and a high number of binding sites available for tracing the biotinylated antibody on the surface. Due to the inherent problems of tracing analyte with a complex of biotinylated antibody and streptavidin-coated nanoparticles, the streptavidin-coated nanoparticles reacting with the surface-captured analyte and biotinylated antibody was favoured and factors influencing this are discussed. This universal labelling technology can be applied to detect any biotinylated molecule, either in solution or on a solid phase, in order to improve detection sensitivities in many areas of biochemical analysis, such as cyto- and histochemistry, multianalyte DNA-chip assays and single-particle assays.

Supersensitive time-resolved immunofluorometric assay of free prostate-specific antigen with nanoparticle label technology.

BACKGROUND: The extreme specific activity of the long-lifetime fluorescent europium(III) chelate nanoparticles and the enhanced monovalent binding affinity of multivalent nanoparticle-antibody bioconjugates are attractive for noncompetitive immunoassay. METHODS: We used a noncompetitive, two-step immunoassay design to measure free prostate-specific antigen (PSA). Europium(III) chelate nanoparticles (107 nm in diameter) were coated with a monoclonal anti-PSA antibody (intrinsic affinity, 6 x 10(9) L/mol). The nanoparticle-antibody bioconjugates had an average of 214 active binding sites per particle and a monovalent binding affinity of 7 x 10(10) L/mol. The assay was performed in a low-fluorescence microtitration well passively coated with an another monoclonal anti-PSA antibody (affinity, 2 x 10(10) L/mol), and the europium(III) fluorescence was measured directly from the bottom of the well by a standard time-resolved microtitration plate fluorometer. RESULTS: The detection limit (mean + 2 SD) was 0.040 ng/L (7.3 x 10(5) molecules/mL), and the dynamic detection range covered four orders of magnitude in a 3-h total assay time. The imprecision (CV) over the whole assay range was 2-10%. The detection limit of the assay was limited by the fractional nonspecific binding of the bioconjugate to the solid phase (0.05%), which was higher than the nonspecific binding of the original antibody (<0.01%). CONCLUSIONS: The sensitivity of the new assay is equal to that of the ambient-analyte, microspot immunoassay and will be improved by use of optimized, high binding-site density nanoparticle-antibody bioconjugates with reduced nonspecific binding and improved monovalent binding affinity.