Mitotic figure recognition: agreement among pathologists and computerized detector

Despite the prognostic importance of mitotic count as one of the components of the Bloom-Richardson grade, several studies have found that pathologists' agreement on the mitotic grade is fairly modest. Collecting a set of more than 4,200 candidate mitotic figures, we evaluate pathologists' agreement on individual figures, and train a computerized system for mitosis detection, comparing its performance to the classifications of three pathologists. The system's and the pathologists' classifications are based on evaluation of digital micrographs of hematoxylin and eosin stained breast tissue. On figures where the majority of pathologists agree on a classification, we compare the performance of the trained system to that of the individual pathologists. We find that the level of agreement of the pathologists ranges from slight to moderate, with strong biases, and that the system performs competitively in rating the ground truth set. This study is a step towards automatic mitosis count to accelerate a pathologist's work and improve reproducibility.     

Automated measurement of MIB-1 positive area as an alternative to counting in follicular lymphoma

Manual counting of MIB-1 positive cells which has been suggested as an alternative to centroblast counting for the diagnostic grading of follicular lymphoma is a laborious task. In this study, the validity of automated measurement of the MIB-1 positive area is analyzed as an alternative approach. Archival MIB-1 stained tissue sections of 15 follicular lymphomas were assessed manually and automatically by three independent observers. Concordance correlation coefficients and coefficients of variation were calculated to study reproducibility and variability of both methods and to compare result from both methods. A good concordance was observed between the two methods. The reproducibility of the automated method was slightly better than the manual counting of positive nuclei. Measurement of MIB-1 positive surface area may be used as a simple and fast alternative to tedious manual counting of positive nuclei as a potential help in follicular lymphoma grading.     

Reliability and intermethod agreement for body fat assessment among two field and two laboratory methods in adolescents

To increase knowledge about reliability and intermethods agreement for body fat (BF) is of interest for assessment, interpretation, and comparison purposes. It was aimed to examine intra- and inter-rater reliability, interday variability, and degree of agreement for BF using air-displacement plethysmography (Bod-Pod), dual-energy X-ray absorptiometry (DXA), bioelectrical impedance analysis (BIA), and skinfold measurements in European adolescents. Fifty-four adolescents (25 females) from Zaragoza and 30 (14 females) from Stockholm, aged 13-17 years participated in this study. Two trained raters in each center assessed BF with Bod-Pod, DXA, BIA, and anthropometry (DXA only in Zaragoza). Intermethod agreement and reliability were studied using a 4-way ANOVA for the same rater on the first day and two additional measurements on a second day, one each rater. Technical error of measurement (TEM) and percentage coefficient of reliability (%R) were also reported. No significant intrarater, inter-rater, or interday effect was observed for %BF for any method in either of the cities. In Zaragoza, %BF was significantly different when measured by Bod-Pod and BIA in comparison with anthropometry and DXA (all P < 0.001). The same result was observed in Stockholm (P < 0.001), except that DXA was not measured. Bod-Pod, DXA, BIA, and anthropometry are reliable for %BF repeated assessment within the same day by the same or different raters or in consecutive days by the same rater. Bod-Pod showed close agreement with BIA as did DXA with anthropometry; however, Bod-Pod and BIA presented higher values of %BF than anthropometry and DXA.     

Limited agreement among three global gene expression methods highlights the requirement for non-global validation

MOTIVATION: DNA microarrays have revolutionized biological research, but their reliability and accuracy have not been extensively evaluated. Thorough testing of microarrays through comparison to dissimilar gene expression methods is necessary in order to determine their accuracy. RESULTS: We have systematically compared three global gene expression methods on all available histologically normal samples from five human organ types. The data included 25 Affymetrix high-density oligonucleotide array experiments, 23 expressed sequence tag based expression (EBE) experiments and 5 SAGE experiments. The reported gene-by-gene expression patterns showed a wide range of correlations between pairs of methods. This level of agreement was sufficient for accurate clustering of datasets from the same tissue and dissimilar methods, but highlights the need for thorough validation of individual gene expression measurements by alternate, non-global methods. Furthermore, analyses of mRNA abundance distributions indicate limitations in the EBE and SAGE methods at both high- and low-expression levels.     

MIB-1 and S-phase cell fraction predict survival in non-Hodgkin's lymphomas

The monoclonal antibody anti-Ki67 is used to detect proliferating cells, but its main limitation is the requirement of fresh-frozen material. On a series of patients with non-Hodgkin's lymphoma, we used a Ki67 equivalent monoclonal antibody, the recently proposed MIB-1, on formalin-fixed histopathological material using microwave antigen retrieval. MIB-1 expression was analysed in relation to other proliferation indices, such as autoradiographic ³H-thymidine labelling index (³HTL1) and flow cytometric S-phase cell fraction (FCM-S) and to pathological status. Moreover, the prognostic relevance of the cell kinetic indices was defined in uni- and multivariate analyses including histology and tumour stage. The relationship between MIB-1 index and the other proliferation indices was statistically significant even though the correlation coefficient was around 0.6. The MIB-1 index was also related to the REAL (Revised European American Lymphoma) classification, but not to the Ann Arbor stage classification. Univariate analysis showed that the MIB-1 index was a significant predictor of 6-year survival in the overall series and in distinctly analysed low-grade and high-grade lymphoma subgroups. With regard to S-phase indices, ³HTLI was a powerful prognosticator in patients with high-grade histologies and FCM-S in patients with low-grade histologies. Multivariate analyses revealed that MIB-1 indiex, ³HTLI and FCM-S retained their prognostic significance independent of histology. In conclusion, the MIB-1 antibody provides prognostic information in non-Hodgkin's lymphomas and has the main advantage that it can be used in formalin-fixed, paraffin-embedded specimens.     

浮游甲壳类定量采样计数方法评价

１引言浮游生物是养殖鱼类特别是滤食性鱼类的主要饵料,同时又是评价水生态系统质量优劣的重要指标．在大水面增养殖研究中,常常通过水体中浮游生物的采样分析,计算水体生物生产力．而在池塘生态系统中则又常常借助于浮游生物的采样分析来判定养鱼水质,决定增施饵肥料...     

A comparison of methods for counting viruses in aquatic systems

In this study, we compared different methods-including transmission electron microscopy-and various nucleic acid labeling methods in which we used the fluorochromes 4',6'-diamidino-2-phenylindole (DAPI), 4-[3-methyl-2,3-dihydro-(benzo-1, 3-oxazole)-2-methylmethyledene]-1-(3'-trimethyl ammoniumpropyl)-quinilinium diioide (YOPRO-1), and SYBR Green I, which can be detected by epifluorescence microscopy (EM), for counting viruses in samples obtained from freshwater ecosystems whose trophic status varied and from a culture of T7 phages. From a quantitative and qualitative viewpoint, our results showed that the greatest efficiency for all ecosystems was obtained when we used the EM counting protocol in which YOPRO-1 was the label, as this fluorochrome exhibited strong and very stable fluorescence. A modification of the original protocol in which YOPRO-1 was used is recommended, because this modification makes the protocol faster and allows it to be used for routine analysis of fixed samples. Because SYBR Green I fades very quickly, the use of this fluorochrome is not recommended for systems in which the viral content is very high (>10(8) particles/ml), such as treated domestic sewage effluents. Experiments in which we used DNase and RNase revealed that the number of viruses determined by EM was slightly overestimated (by approximately 15%) because of interference caused by the presence of free nucleic acids.     

Spike-frequency counting and integrative methods for analysis of aortic nerve activity

1. -- Aortic nerve activity was simultaneously measured in anaesthetized rabbits by spike-frequency counting and integrative methods during decreases by bleeding in mean blood pressure (MBP) from 130 to 50 mm Hg. Values of counted and integrated activities were expressed in arbitrary units. 2. -- No statistically significant difference between the counted and integrated activity was pointed out in the range of MBP from 130 to 90 mm Hg. 3. -- Below a MBP of 90 mm Hg, the counted activity was significantly higher than the integrated activity. This difference could be the result of a better appraisal of the number of small and medium spikes by the counting equipment than by the integrating system.     

Comparison of fluorescence microscopy and solid-phase cytometry methods for counting bacteria in water

Total direct counts of bacterial abundance are central in assessing the biomass and bacteriological quality of water in ecological and industrial applications. Several factors have been identified that contribute to the variability in bacterial abundance counts when using fluorescent microscopy, the most significant of which is retaining an adequate number of cells per filter to ensure an acceptable level of statistical confidence in the resulting data. Previous studies that have assessed the components of total-direct-count methods that contribute to this variance have attempted to maintain a bacterial cell abundance value per filter of approximately 10(6) cells filter(-1). In this study we have established the lower limit for the number of bacterial cells per filter at which the statistical reliability of the abundance estimate is no longer acceptable. Our results indicate that when the numbers of bacterial cells per filter were progressively reduced below 10(5), the microscopic methods increasingly overestimated the true bacterial abundance (range, 15.0 to 99.3%). The solid-phase cytometer only slightly overestimated the true bacterial abundances and was more consistent over the same range of bacterial abundances per filter (range, 8.9 to 12.5%). The solid-phase cytometer method for conducting total direct counts of bacteria was less biased and performed significantly better than any of the microscope methods. It was also found that microscopic count data from counting 5 fields on three separate filters were statistically equivalent to data from counting 20 fields on a single filter.     

Thyroid cell proliferation in Graves' disease. Use of MIB-1 monoclonal antibody

OBJECTIVE: To measure thyroid cell proliferation in patients with Graves' disease (GD) before and during treatment with antithyroid drugs. STUDY DESIGN: Patients were assessed by fine needle aspiration biopsy before (n = 20) and after 4 (n = 19) and 12 months of treatment (n = 15) with propylthiouracil or methimazole. Cell proliferation index (CPI) was estimated by immunocytochemistry using MIB-1. CPI was studied in relation to the cytologic parameters of the smears; clinical parameters, such as Wayne's Clinical Index (WCI) and time without treatment; laboratory parameters, such as 131I uptake and dosage of serum free thyroxin and thyroid-stimulating hormone; and thyroid ultrasound. RESULTS: CPI varied from 0.00% to 25.00% before treatment, 0.00% to 23.00% at 4 months and 0.00% to 14.84% at 12 months. CPI median values were 6.50%, 4.30% and 3.30%, respectively (before and after 4 months and 12 months of treatment). CPI had a positive correlation with WCI and FT4 at 12 months of treatment. CONCLUSION: Thyroid CPI in GD varies from case to case. However, due to its decreasing pattern during follow-up and its positive correlation with thyrotoxicosis severity, CPI may indicate the functional status of the gland and contribute to a better understanding of GD.     

MIB-1 immunohistometry of follicular adenoma and follicular carcinoma of the thyroid gland

OBJECTIVE: To analyze cellular proliferative activity, MIB-1 immunopositivity of normal tissue (n = 20), follicular adenoma (n = 30) and follicular carcinoma (n = 32) of the thyroid gland was analyzed by means of immunohistometry. STUDY DESIGN: Immunohistochemical reactions were performed on 3-micron sections from routinely formalin fixed and paraffin embedded surgical specimens using an indirect peroxidase method. The rate of immunostained cells was determined using the CM-2 TV image analysis system (Hund, Wetzlar, Federal Republic of Germany). Forty viewing fields (1.94 mm2) were measured with 20:1 objective magnification. An average of 5,965 cells were assessed in each case. RESULTS: Mean MIB-1 immunopositivity was higher in follicular carcinoma (average, 2.30%) and follicular adenoma (0.58%) than in normal thyroid tissue (0.14%). The distribution of single values differed significantly between groups (P < .001). To test the suitability of MIB-1 immunohistometry for the differential diagnosis of follicular adenoma and follicular carcinoma, different four-field tables with varying thresholds were calculated. Using a threshold of 0.9%, follicular carcinoma could be detected with a sensitivity of 75% (24/32) and a specificity of 83% (25/30). If a specificity of 90% is required (27/30), the sensitivity of the test decreases to 69% (22/32), based on a threshold of 1.1%. CONCLUSION: As some overlap of single values has to be considered, MIB-1 immunohistometry, although presenting new insights into the proliferative potential of thyroid lesions, is of only limited value for the differential diagnosis of follicular lesions in routine surgical pathology.     

Quench correction in Cerenkov counting: channels ration and external source channels ratio methods

Channels ratio and external source ratio methods which may be used for accurate quench correction in ?erenkov counting systems have been developed. In general, the channels ratio method will be more accurate for quench correction in high count rate samples where good ratio statistics are available. For low level samples, the external source channels ratio method will provide more accurate quench correction if efficiencies are greater that 10%.     

Reinke's Edema: investigations on the role of MIB-1 and hepatocyte growth factor

Reinke''s edema is a benign disease of the human vocal fold, which mainly affects the sub-epithelial layer of the vocal fold. Microscopic observations show a strongly oedematous epithelium with loosened intercellular junctions, a disruption of the extracellular connections between mucosal epithelium and connective tissue, closely adherent to the thyroarytenoid muscle. Thickening of the basal layer of epithelium, known as Reinke''s space, high deposition of fibronectin and chronic inflammatory infiltration it is also visible. We analyzed, together with the hepatocyte growth factor (HGF), the expression level of MIB-1 in samples harvested from patients affected by Reinke''s edema, in order to define its biological role and consider it as a possible prognostic factor in the follow-up after surgical treatment. We observed a moderate expression of HGF in the lamina propria of the human vocal fold and in the basal membrane of the mucosal epithelium. Our finding suggests that this growth factor acts as an anti - fibrotic agent in Reinke''s space and affects the fibronectin deposition in the lamina propria. MIB-1, on the contrary, showed a weak expression in the basement membrane of the mucosal epithelium and a total absence in the lamina propria deep layer, thus suggesting that only the superficial layer is actively involved in the reparatory process with a high regenerative capacity, together with a high deposition of fibronectin. The latter is necessary for the cellular connections reconstruction, after the inflammatory infiltration.Key words: Reinke''s edema, Ki67, HGF, fibronectin, immunohistochemistry, extracellular matrix.     

Immunoreactivity to MIB-1 in breast cancer: methodological assessment and comparison with other proliferation indices

The recent availability of the monoclonal antibody MIB-1 (which is able to detect the human nuclear cell proliferation-associated antigen Ki-67 even on formalin-fixed, paraffin-embedded sections, microwave-treated and routinely processed for immunohistochemistry) could open new avenues for validation of the clinical role of tumour cell proliferation on large, consecutive and unselected series of human tumours. However, the routine use of such a marker requires a methodological standardization as well as the comparative assessment of some technical and biological aspects. The MIB-1 index was determined in parallel samples from 50 consecutive invasive breast carcinomas processed with different fixatives for different times. The median values of MIB-1 indices following 2, 6 and 24 h of formalin fixation were similar (29.4%, 30.6% and 29.7%, respectively) and consistent with those reported in the literature; squared linear regression coefficients were 0.99. The median values of MIB-1 indices were markedly lower in Bouin-fixed, paraffin-embedded, and in frozen samples (20.0% and 19.8%, respectively), with a poor correlation coefficient with the values detected following formalin fixation ( R ²=0.456). Moderate and poor correlations were observed between Ki-67 index and MIB-1 detected on frozen ( r _s, 0.78) or formalin-fixed, paraffin-embedded samples ( r _s, 0.47) and a minimal concordance was observed between TLI and MIB-1 or Ki-67 ( r _s, 0.25 and 0.22, respectively). Our results indicate interference of the fixative type on immunoreactivity to MIB-1 and also suggest that Ki-67 and MIB-1 reacted with different epitopes of the same antigen.