Treatment of multiple adjacent RT 1 gingival recessions with the modified coronally advanced tunnel (MCAT) technique and a collagen matrix or palatal connective tissue graft: 9-year results of a split-mouth randomized clinical trial.

OBJECTIVES: To evaluate t he long-term outcomes following treatment of RT 1 multiple adjacent gingival recessions (MAGR) using the modified coronally advanced tunnel (MCAT) with either a collagen matrix CM or a connective tissue graft (CTG). MATERIAL AND METHODS: Sixteen of the original 22 subjects included in a randomized, controlled split-mouth clinical trial were available for the 9-year follow-up (114 sites). Recessions were randomly treated by means of MCAT + CM (test) or MCAT + CTG (control). Complete root coverage (CRC), mean root coverage (MRC), gingival recession depth (GRD), probing pocket depth (PD), keratinized tissue width (KTW), and thickness (KGT) were compared with baseline values and with the 12-month results. RESULTS: After 9 years, CRC was observed in 2 patients, one in each group. At 9 years, MRC was 23.0 ± 44.5% in the test and 39.7 ± 35.1% in the control group (p = 0.179). The MRC reduction compared to 12 months was - 50.1 ± 47.0% and - 48.3 ± 37.7%, respectively. The upper jaw obtained 31.92 ± 43.0% of MRC for the test and 51.1 ± 27.8% for the control group (p = 0.111) compared to the lower jaw with 8.3 ± 46.9% and 20.7 ± 40.3%. KTW and KGT increased for both CM and CTG together from 2.0 ± 0.7 to 3.1 ± 1.0 mm (< 0.0001). There were no statistically significant changes in PD. CONCLUSION: The present results indicate that (a) treatment of MAGR using MCAT in conjunction with either CM or CTG is likely to show a relapse over a period of 9 years, and (b) the outcomes obtained in maxillary areas seem to be more stable compared to the mandibular ones. CLINICAL RELEVANCE: The mean root coverage at 12 months could not be fully maintained over 9 years. On a long-term basis, the results seem to be less stable in the mandible as compared to maxillary areas.

Plastic waste from marine environment: Demonstration of possible routes for recycling by different manufacturing technologies.

The increasing amount of marine plastic waste poses challenges including, not only the collection, but also the subsequent recyclability of the plastic. An artificial accelerated weathering procedure was developed, which modelled the marine environment and investigated the recyclability of weathered and non-weathered PET. Marine conditions were simulated for poly(ethylene terephthalate) (PET) bottle material and high-density polyethylene (HDPE) cap material. It consisted of 2520 h cyclical weathering, alternating the sample between a salt spray and a Xenon-chamber-this corresponds to roughly 3-4 years on the surface of an ocean. It was proved that the molecular weight of PET is a function of weathering time and can be described mathematically. Microscopic examination of the surface of the PET bottles and HDPE caps proved that these surfaces were damaged. After weathering, manufacturing tests were performed on the PET material by extrusion, injection moulding, 3D printing and thermoforming. Quantitative comparison between products manufactured by the same technology was performed in order to compare the qualities of products made from original PET, non-weathered PET waste, which was the example of classical recycling, and weathered PET. In the case of products made from weathered PET, certain mechanical and optical properties (e.g. impact strength and transparency) were significantly impaired compared to the original PET and the recycled, non-weathered PET. Certain other properties (e.g. strength and rigidity) did not change significantly. It was proved that the samples from weathered plastic material can be successfully recycled mechanically and used to manufacture plastic products.

Correction to: Abstracts : 31st European Congress of Pathology.

The name of the co-author of MD-05-001 (page S62) should be presented as 'S. Vári-Kakas' instead of 'I.î Vári-Kakas' in the authorship group. The name has been corrected in the authorship group shown above.

Analysis of microRNA expression in brush cytology specimens improves the diagnosis of pancreatobiliary cancer.

BACKGROUND/OBJECTIVES: Malignant pancreatobiliary strictures are in many cases clinically indistinguishable and present a major problem to endoscopy specialists. Intraductal sampling procedures such as brush cytology are commonly used for diagnosis with a sensitivity that is low for a diagnostic test used in daily clinical practice. MicroRNA (miR) alterations detected in many cancers are disease-specific, which can be utilized in clinical applications. The aim of the present study was to analyze whether determination of miR expression levels in intraductal brush cytology specimens is a feasible approach to improve the diagnosis of pancreatobiliary cancer. METHODS: Brush cytology specimens have been collected during endoscopic retrograde cholangio-pancreatography (ERCP) and analyzed by routine cytology and ancillary miR assays. Total RNA was extracted using the miRNeasy Mini Kit and the expression of miRs frequently dysregulated in pancreatobiliary cancer (miR-16, miR-21, miR-196a, miR-221) were analyzed by quantitative real-time PCR using RNU6B as internal control. RESULTS: Routine cytology resulted in no false positive diagnoses, however, the combined sensitivity remained at 53.8%. Expression (ΔCt values) of miR-16 (p = 0.0039), miR-196a (p = 0.0003) and miR-221 (p = 0.0049) showed a clear statistical significance between malignant and benign pancreatobiliary specimens (n = 35). Malignancy could be detected combining routine cytology and the miR-196a single marker expression levels with a sensitivity of 84.6% (92.9% in biliary strictures) with no false positives. CONCLUSIONS: The results offer the first direct demonstration that microRNAs are readily detectable in brush cytology specimens obtained during ERCP, and have the potential to help the cytological diagnosis of pancreatobiliary malignancy.

Alteration of sarcoplasmic reticulum Ca2+ ATPase expression in lower limb ischemia caused by atherosclerosis obliterans.

Atherosclerosis is a disease caused by a build-up of fatty plaques and cholesterol in the arteries. The lumen of the vessels is obliterated resulting in restricted blood supply to tissues. In ischemic conditions, the cytosolic Ca2+ level of skeletal muscle may increase, indicating the alteration of Ca2+ removal mechanisms. Ca2+ is transported from cytosol into the sarcoplasmic reticulum by Ca2+ ATPase (SERCA), with its 1a isoform expressed in adult, while its 1b isoform in neonatal and regenerating fast-twitch skeletal muscle. To investigate the role of these isoforms in ischemic skeletal muscle, biopsies from musculus biceps femoris of patients who underwent amputation due to atherosclerosis were examined. Samples were removed from the visibly healthy and hypoxia-affected tissue. Significantly increased SERCA1a expression was detected under the ischemic conditions (246 ± 69%; p < 0.05) compared with the healthy tissue. Furthermore, the ratio of SERCA1a-positive fibers was slightly increased (46 ± 4% in healthy tissue and 60 ± 5% in ischemic tissue; p > 0.05), whereas SERCA2a did not change. In addition, in primary cultures derived from hypoxia-affected tissue, the diameter and fusion index of myotubes were significantly increased (30 ± 1.6 µm vs. 41 ± 2.4 µm and 31 ± 4% vs. 45 ± 3%; p < 0.05). We propose that the increased SERCA1a expression indicates the existence and location of compensating mechanisms in ischemic muscle.

Molecular cloning, mRNA expression and biological activity of the pheromone biosynthesis activating neuropeptide (PBAN) from the European corn borer, Ostrinia nubilalis.

Pheromone biosynthesis activating neuropeptide (PBAN) is a member of the pyrokinin (FXPRLamide) insect neuropeptides. Here, we report the cloning of the gene Ostnu-PBAN from the E and Z pheromone strains of the European corn borer (ECB), Ostrinia nubilalis (Lepidoptera: Crambidae), a major pest of maize. The Ostnu-PBAN genomic sequence is > 5 kb in length and consists of six exons. The deduced amino acid sequence revealed a 200-residue precursor protein including a signal peptide, a 24-amino acid (aa) diapause hormone, a 37-aa PBAN and three other FXPRLamide neuropeptides. Our in vivo assays suggest that the 37-aa synthetic Ostnu-PBAN is hormonally active in the pheromone gland. It restores sex pheromone production to normal levels in mated females and decapitated virgins of both E and Z cultures. The results of a real-time PCR analysis indicated that Ostnu-PBAN mRNA levels reached a plateau in the brain-suboesophageal ganglion complexes 1 day after eclosion, and mating did not affect the mRNA expression. Three size classes of Ostnu-PBAN mRNA (1.9, 2.0 and 2.1 kb) were obtained, differing only in the length of the 3' untranslated region. However, there was no correlation between sequence divergence and the pheromone composition, voltinism or geographical origin (Hungary, Slovenia, Sweden, Turkey) of ECB moths.

Organizer and regulatory role of colonic isolated lymphoid follicles in inflammation.

Gut-associated lymphoid tissue (GALT) is supposed to play an integral role in the organization of colonic repair mechanisms. Majority of the GALT is composed of isolated and aggregated lymphoid follicles distributed throughout the intestines. These lymphoid follicles, including Peyer's patches of the small, and isolated lymphoid follicles (ILFs) of both the small and large intestines, are composed of a specialised follicle associated epithelium overlying a subepithelial dome containing numerous dendritic cells, macrophages, T and B cells. Within inflammatory conditions the number, the diameter and the density of ILFs are increasing. Follicles are involved not just in immune surveillance, but their presence is also indispensable for normal colonic mucosal regeneration. Regarding mucosal repair the relation of ILFs to bone marrow derived stem cells, follicular dendritic cells, subepithelial myofibroblasts and crypt formations, and the putative organizer role of ILFs have not been clarified yet.

Volumetric difference evaluation of registered three-dimensional pre-operative and post-operative CT dental data.

OBJECTIVE: The purpose of this study is to propose a complete methodology for automatically registering three-dimensional (3D) pre-operative and post-operative CT scan dental volumes as well as to provide a toolset for quantifying and evaluating their volumetric differences. METHODS: The proposed methodology was applied to cone beam CT (CBCT) data from 20 patients in order to assess the volume of augmented bone in the alveolar region. In each case, the pre-operative and post-operative data were registered using a 3D affine-based scheme. The performance of the 3D registration algorithm was evaluated by measuring the average distance between the edges of the registered sets. The differences between the registered sets were assessed through 3D subtraction radiography. The volume of the differences was finally evaluated by defining regions of interest in each slice of the subtracted 3D data and by combining all respective slices to model the desired volume of interest. The effectiveness of the algorithm was verified by applying it to several reference standard-shaped objects with known volumes. RESULTS: Satisfactory alignment was achieved as a low average offset of 1.483 ± 1.558 mm was recorded between the edges of the registered sets. Moreover, the estimated volumes closely matched the volumes of the reference objects used for verification, as the recorded volume differences were less than 0.4 mm(3) in all cases. CONCLUSION: The proposed method allows for automatic registration of 3D CBCT data sets and the volumetric assessment of their differences in particular areas of interest. The proposed approach provides accurate volumetric measurements in three dimensions, requiring minimal user interaction.

Is blood thicker than water? Social support, depression and the modifying role of ethnicity/nativity status.

BACKGROUND: Social support may be particularly important in countering depression among systematically disadvantaged groups. Latino immigrants are an example of a disadvantaged population that has better than expected mental health outcomes. One explanation put forth for this pattern is strong social support from kin networks. Studies on the effect of social support on mental health often assess the quantity of social ties rather than the quality of the support they provide. In addition, such studies rarely specify the source of support and how support from family versus friends may differentially impact mental health. METHODS: In this study, data from the Project on Human Development in Chicago Neighbourhoods were used to disaggregate the effects of source-specific emotional support on risk of depression. Second, the relationship between ethnicity/nativity status and risk of depression was examined. Finally, whether the relationship between family-based and friend-based social support and depression differed across ethnic/nativity status was explored. RESULTS: Support from both family and friends had protective effects on risk of depression; however, when mutually adjusted, only kin support remained statistically significant. At higher levels of family support, foreign-born Mexicans and African Americans had decreased risk of depression than at low levels of family support. CONCLUSION: This study provides evidence that family support may be more important than non-kin support for mental health. Findings also suggest that the effects of family support on risk of depression vary by ethnicity and nativity status. Preservation of naturally occurring support resources among some groups may be a way to maintain mental health.

Reversal of gene expression changes in the colorectal normal-adenoma pathway by NS398 selective COX2 inhibitor.

BACKGROUND AND AIMS: Treatment of colorectal adenomas with selective cyclooxygenase-2 inhibitors can contribute to the chemoprevention of colorectal cancer (CRC), but the molecular background of their effect is not fully understood. We analysed the gene expression modulatory effect of N-(2-cyclohexyloxy-4-nitrophenyl)-methanesulfonamide (NS398) on HT29 cells to be correlated with expression data gained from biopsy samples. METHODS: HT29 colon adenocarcinoma cells were treated with NS398, and global mRNA expression was analysed on HGU133Plus2.0 microarrays. Discriminatory transcripts between normal and adenoma and between adenoma and CRC biopsy samples were identified using HGU133Plus2.0 microarrays. The results were validated using RT-PCR and immunohistochemistry. RESULTS: Between normal and adenoma samples, 20 classifiers were identified, including overexpressed cadherin 3, KIAA1199, and downregulated peptide YY, glucagon, claudin 8. Seventeen of them changed in a reverse manner in HT29 cells under NS398 treatment, 14 (including upregulated claudin 8, peptide YY, and downregulated cadherin 3, KIAA1199) at a significance of P<0.05. Normal and CRC could be distinguished using 38 genes, the expression of 12 of them was changed in a reverse manner under NS398 treatment. CONCLUSION: NS398 has a reversal effect on the expression of several genes that altered in colorectal adenoma-carcinoma sequence. NS398 more efficiently inverted the expression changes seen in the normal-adenoma than in the normal-carcinoma transition.

Recent psychopathology, suicidal thoughts and suicide attempts in households with and without firearms: findings from the National Comorbidity Study Replication.

OBJECTIVE: To assess the relationship between firearm ownership and possible psychiatric confounders of the firearm-suicide relationship. METHODS: Multivariate logistic regression was used to estimate the association between living in a home with firearms and 12-month occurrence of major Diagnostic and statistical manual of mental disorders (DSM)-IV disorders and suicidal behaviour among respondents to the National Comorbidity Survey Replication, a household survey of 9282 adults aged 18+. Analyses controlled for sociodemographic characteristics including age, sex, race/ethnicity, educational attainment and poverty. RESULTS: Approximately one in three Americans reported living in a home with firearms. People living in a home with firearms were no more or less likely than people in homes without firearms to have recent (past year) anxiety disorders (OR = 1.0, 95% CI 0.8 to 1.2), mood disorders (OR = 0.9, 95% CI 0.7 to 1.1) or substance dependence and/or abuse (OR = 0.9, 95% CI 0.6 to 1.3). Past year suicidal ideation (OR = 0.8, 95% CI 0.5 to 1.3) and suicide planning (OR = 0.5, 95% CI 0.2 to 1.4) were also not associated with living in households with firearms. Having made a suicide attempt over the previous year was the only outcome more common among participants reporting that they currently lived in a home without [corrected] firearms. CONCLUSIONS: The previously reported association between household firearm ownership and heightened risk of suicide is not explained by a higher risk of psychopathology among gun-owning families. As there are Americans with suicidal ideation and/or significant and recent psychiatric disorders currently living in homes with firearms, future work should focus on understanding the impediments to effectively communicating the suicide risk associated with household firearms.

The potential of digital microscopy in breast pathology.

The rapidly evolving field of digital microscopy supports the efficient exploitation of inherent information from stained glass slides to offer widespread utilization in breast histopathology. Digital image signals can be accurately measured, integrated into databases and shared through computer networks. Therefore, digital microscopy can boost telepathology-consultation, gradual- and postgradual teaching, proficiency testing, intra- and interlaboratory validation of biomarker screening interpretation, and automated image analysis of biomarker expression for both diagnostics and research applications. This is a brief highlight of the potential of digital microscopy in breast pathology applications.

The RR genotype of codon 72 of p53 gene reduces the development of intestinal metaplasia.

BACKGROUND AND AIMS: Mutations of p53 gene can contribute to the development of gastric cancer. Our aims were to evaluate the premalignant gastric intestinal metaplasia-related p53 alterations, using and comparing capillary sequencing and p53 resequencing chip in gastric biopsy and peripheral blood samples. Furthermore we examined the effect of p53 polymorphism on the protein expression level. PATIENTS AND METHODS: Deoxyribonucleic acid was extracted from antral gastric biopsy samples of 50 intestinal metaplasia patients (27 Helicobacter pylori positive, 23 H. pylori negative) and 51 controls (all H. pylori negative). Exon 4 of p53 gene was examined by capillary sequencing (CS). From 7 intestinal metaplasia patients extra deoxyribonucleic acid samples were extracted from blood and from the corpus and from the antrum of the stomach and 5 additional exons were examined by CS and 10 with GeneChip p53 Assay (Affymetrix). In 19 patients p53 immunohistochemistry was performed. RESULTS: RR genotype on codon 72 was found to significantly (p=0.0087) reduce the chance of intestinal metaplasia in H. pylori positive patients as compared to the normal controls. The p53 alterations were identical in antral, corpus and blood samples. The p53 protein expression was in significant correlation with the genetic alterations. CS and chip method-based sequencing results were not in correlation. CONCLUSIONS: According to our results RR genotype decreases the incidence of IM. The genetic background is reflected in the expression of p53 protein. Chip method-based deoxyribonucleic acid sequence data need careful confirmation.

Differentiation potential of stem cells from human dental origin - promise for tissue engineering.

Recent studies have revealed the existence of stem cells in various human tissues including dental structures. We aimed to establish primary cell cultures from human dental pulp and periodontal ligament, to identify multipotential adult stem cells in these cultures, and to study the differentiation capacity of these cells to osteogenic and to neuronal fates. Dental pulp and the periodontal ligament were isolated from extracted human wisdom teeth. The extracellular matrix was enzymatically degraded to obtain isolated cells for culturing. Both dental pulp and periodontal ligament derived cultures showed high proliferative capacity and contained a cell population expressing the STRO-1 mesenchymal stem cell marker. Osteogenic induction by pharmacological stimulation resulted in mineralized differentiation as shown by Alizarin red staining in both cultures. When already described standard neurodifferentiation protocols were used, cultures exhibited only transient neurodifferentiation followed by either redifferentiation into a fibroblast-like phenotype or massive cell death. Our new three-step neurodifferentiation protocol consisting of (1) epigenetic reprogramming, then (2) simultaneous PKC/PKA activation, followed by (3) incubation in a neurotrophic medium resulted in robust neurodifferentiation in both pulp and periodontal ligament cultures shown by cell morphology, immunocytochemistry and real time PCR for vimentin and neuron-specific enolase. In conclusion, we report the isolation, culture and characterization of stem cell containing cultures from both human dental pulp and periodontal ligament. Furthermore, our data clearly show that both cultures differentiate into mineralized cells or to a neuronal fate in response to appropriate pharmacological stimuli. Therefore, these cells have high potential to serve as resources for tissue engineering not only for dental or bone reconstruction, but also for neuroregenerative treatments.

Proton pump inhibitor co-therapy normalizes the increased cell turnover of the gastric mucosa both in NSAID and selective COX-2 users.

Proton pump inhibitor (PPI) co-therapy is considered the best strategy in preventing gastrointestinal complications during non-steroidal anti-inflammatory drug (NSAID) treatment, but there is limited information available on its effect on gastric mucosal cell kinetics. To evaluate the effect of PPI co-therapy on gastric mucosa we investigated epithelial cell proliferation, apoptosis, epithelial growth factor receptor (EGFR) and p53 expression in patients on chronic non-selective NSAID or cyclooxygenase-2 selective inhibitor (COX-2) treatment. Gastric biopsies of the antrum were taken from 10-10 patients on chronic NSAID and COX-2, therapy prior and after 6 months PPI co-therapy, and 10 controls without any treatment. Cell proliferation, apoptosis, EGFR and p53 expression were measured by immunohistochemistry. At least 600 glandular epithel cells were encountered and results were expressed as % of total cells counted. We found increased cell proliferation in patients on chronic COX-2 but not on NSAID therapy. Patients on either NSAID or COX-2 therapy had an increased p53 and decreased EGFR expression. PPI therapy reversed not only the increased cell proliferation and p53 expression, but also the suppressed EGFR expression when administered as co-therapy. The fewer gastrointestinal side effects observed during chronic COX-2 therapy may partially be the result of the higher cell proliferation. This effect is not mediated by the EGFR pathway. PPI co-therapy normalizes the disturbed cell kinetics irrespective of NSAID treatment used.

Review article: Deoxyribonucleic acid-based diagnostic techniques to detect Helicobacter pylori.

Helicobacter pylori is an important cause of many gastrointestinal disorders, ranging from chronic gastritis to gastric lymphoma and adenocarcinoma. The deoxyribonucleic acid-based assays have the potential to be a powerful diagnostic tool given its ability to specifically identify H. pylori deoxyribonucleic acid. Markers used to include general H. pylori structures and pathogenetic factors like ureaseA, cagA, vacA, iceA. Deoxyribonucleic acid or bacterial ribonucleic acid for polymerase chain reaction assays can be collected from gastric biopsy, gastric juice, stool, buccal specimens. Polymerase chain reaction can yield quantitative and genotyping results with sensitivity and specificity that approaches 100%. A clear trend in the direction of the determination of quantitative H. pylori infection by real-time polymerase chain reaction can be observed. Fluorescent in situ hybridization is suggested for routine antibiotic resistance determination. To identify the organism, deoxyribonucleic acid structure and its virulence factors may be feasible by using oligonucleotide microarray specifically recognizing and discriminating bacterial deoxyribonucleic acid and various virulence factors. Deoxyribonucleic acid-based H. pylori diagnosis yields higher sensitivity, however, specificity requires sophisticated labour environment and associated with higher costs.

COX-2-selective inhibitors (COXIBs): gastrointestinal safety.

COX-2 selective inhibitors (coxibs) have been developed with the primary aim to reduce/avoid gastrointestinal (GI) toxicity observed during conventional (non-selective) non-steroidal anti-inflammatory (NSAID) therapy. Coxibs have clearly and convincingly been shown to be superior to conventional NSAIDs with significantly less GI side effects. When hard endpoints such as perforation, obstruction, and serious bleeding considered, coxibs reduce the risk by approximately 50 percent. Although selective COX-2 inhibition seems not to be enough for complete elimination of GI toxicity, coxibs posses no more GI toxicity than placebo in prospective clinical studies and further increase in COX-2 selectivity does not reduce GI toxicity. For the initial aim developed, thus coxibs fulfilled their promise and will soon replace conventional NSAIDs.

Dual role of serum soluble E-cadherin as a biological marker of metastatic development in gastric cancer.

BACKGROUND: Soluble E-cadherin serum levels as a potential biological marker for gastric cancer were analysed with special consideration to clinical and pathological features. METHODS: Seventy-one healthy control subjects and 166 patients with gastric cancer were enrolled. Gastric cancer patients were classified into intestinal-type (51%) and diffuse-type (49%), according to Laurén. Soluble E-cadherin serum levels were measured with enzyme-linked immunosorbent assay. RESULTS: The mean logarithmic concentrations of soluble E-cadherin in gastric cancer patients were significantly higher than those of control subjects, with an average of 4.03 (+/- 0.32) versus 3.86 (+/- 0.24), respectively (P < 0.0001). The concentration of soluble E-cadherin was significantly higher in the intestinal-type group than in the diffuse-type group, with an average of 4.07 +/- 0.3 versus 3.98 +/- 0.34, respectively (P = 0.0494). In the intestinal-type group, concentrations of soluble E-cadherin were significantly higher in more advanced stages (stages III-IV) than in earlier stages (stages I-II), with an average of 4.13 +/- 0.29 versus 3.96 +/- 0.31, respectively (P = 0.0234). In the diffuse-type group, concentrations of soluble E-cadherin were significantly higher in localized than in metastatic gastric cancer, with an average soluble E-cadherin concentation of 4.15 +/- 0.3 versus 3.95 +/- 0.32, respectively (P = 0.0139). CONCLUSION: Serum soluble E-cadherin concentrations exhibit a completely different pattern in intestinal-type and diffuse-typegastric cancer. Serum levels are increased in intestinal-type gastric cancer, especially in advanced stages, whereas in diffuse-type gastric cancer E-cadherin levels are decreased in advanced, metastasized cancer.We conclude that soluble E-cadherin concentrations should be interpreted along with Laurén classification and thus might serve as a biological marker in intestinal-type gastric cancer.

Digital slide and virtual microscopy based routine and telepathology evaluation of routine gastrointestinal biopsy specimens.

AIMS: To evaluate a recently developed digital slide and virtual microscope system, and to compare this method with optical microscopy on routine gastrointestinal biopsy specimens in both local and remote access modes. METHODS: A fully computer controlled commercial microscope was used. The scanning program included object detection, autofocus, and image compression algorithms. The overall hard disk space for a gastric biopsy was between 30 and 50 MB and the scanning time was between 20 and 40 minutes. Haematoxylin and eosin stained routine gastric (61) and colon (42) biopsy specimens were selected, scanned, and evaluated by two specialists on an optical (OM) and virtual microscope (VM). RESULTS: The overall concordance of VM and OM with the consensus diagnosis was 95.1% and 97%, respectively. Clinically important concordance was 96.1% and 98% for VM and OM, respectively. The two methods showed concordance in 92% of cases and clinically important concordance in 94.1% of cases. The reasons for discordance were image quality (one case), interpretation difference (three cases), and insufficient clinical information (three cases). Remote evaluation of the digital slides through the Internet has the advantages of the previously used static and dynamic telepathology methods. CONCLUSIONS: Diagnostic concordance was found between OM and VM. The digital slide and the virtual microscope can be alternative techniques in the computerisation of the histology laboratory and in teleconsultation services after further evaluation of time and storage constraints.

Reliability of quantitative reverse-transcriptase-PCR-based detection of tumour cells in the blood between different laboratories using a standardised protocol.

Differences in methods of reverse-transcriptase (RT)-polymerase chain reaction (PCR)-based detection of tumour cells in the blood gives rise to conflicting results, and standardisation is urgently needed. This pilot study aimed to assess the variation of RT-PCR-based detection of tumour cells in blood between four different laboratories using a commercially available kit with a standardised protocol. This kit allows comparison of results from different laboratories and facilitates the investigation of the influence of pre-analytical parameters. All laboratories analysed identical sets of blood samples spiked with tumour cells in a concentration range of 1-100 tumour cells/ml. To study at which level variation was introduced, three kinds of sample sets were generated in which (i) tumour cell RNA was spiked in the RNA of mononuclear cells (MNC), (ii) tumour cells were spiked in isolated MNC, and (iii) tumour cells were spiked in blood. Real-time quantitative RT-PCR was used to detect and quantify cytokeratin 20 (CK20) expression, which is indicative for the presence of epithelial tumour cells. All laboratories were able to detect CK20 expression in all spiked-RNA samples with limited variation in expression levels between laboratories. There was a positive correlation between the amount of spiked tumour cell RNA and CK20 expression level. RT-PCR analysis of spiked-MNC samples resulted in more variation in the CK20 expression levels between laboratories, however again all spiked samples were reported to be positive by all of the laboratories. The evaluation of spiked-blood samples gave rise to considerable quantitative and qualitative variation between the laboratories. Our results underline the importance and need for standardisation and extended quality control studies in the field of pre-analytics.