Taking molecular pathology to the next level: Whole slide multicolor confocal imaging with the Pannoramic Confocal digital pathology scanner.

The emergence and fast advance of digital pathology allows the acquisition, digital storage, interactive recall and analysis of morphology at the tissue level. When applying immunohistochemistry, it also affords the correlation of morphology with the expression of one or two specific molecule of interest. The rise of fluorescence pathology scanners expands the number of detected molecules based on multiplex labeling. The Pannoramic Confocal (created by 3DHistech, Hungary) is a first-of-the-kind digital pathology scanner that affords not only multiplexed fluorescent detection on top of conventional transmission imaging, but also confocality. We have benchmarked this scanner in terms of stability, precision, light efficiency, linearity and sensitivity. X-Y stability and relocalisation precision were well below resolution limit (≤50 nm). Light throughput in confocal mode was 4-5 times higher than that of a point scanning confocal microscope, yielding similar calculated confocal intensities but with the potential for improving signal to noise ratio or scan speed. Response was linear with R2 ≥ 0.9996. Calibrated measurements showed that using indirect labeling ≥2000 molecules per cell could be well detected and imaged on the cell surface. Both standard-based and statistical post-acquisition flatfield corrections are implemented. We have also measured the point spread function (PSF) of the instrument. The dimensions of the PSF are somewhat larger and less symmetric than of the theoretical PSF of a conventional CLSM, however, the spatial homogeneity of these parameters allows for obtaining a specific system PSF for each optical path and using it for optional on-the-fly deconvolution. In conclusion, the Pannoramic Confocal provides sensitive, quantitative widefield and confocal detection of multiplexed fluorescence signals, with optical sectioning and 3D reconstruction, in addition to brightfield transmission imaging. High speed scanning of large samples, analysis of tissue heterogeneity, and detection of rare events open up new ways for quantitatively analyzing tissue sections, organoid cultures or large numbers of adherent cells.

Comparison of virtual microscopy and real microscopy for learning oral histology laboratory course among dental students.

Background/purpose: Virtual microscopy has been used for learning and teaching histology and pathology laboratory course for more than 10 years. This study aimed to compare the learning outcome of virtual microscopy with that of real light microscopy in oral histology laboratory course among dental students. Materials and methods: The study included twenty-three undergraduate dental students who took the oral histology laboratory course. The evaluation methods of the study were divided into two parts: questionnaire survey (6 questions) and histological diagnosis ability assessment tests. The data were collected and analyzed statistically. Results: Dental students showed a significantly higher acceptance rate of virtual microscopy than that of real light microscopy for learning oral histology laboratory course (all P-values < 0.01). Moreover, dental students using virtual microscopy with digitized virtual slides for learning oral histology laboratory course possessed significantly better histological diagnosis ability than those using real light microscopy with traditional glass slides for learning oral histology laboratory course (P < 0.001). Conclusion: Virtual microscopy with digitized virtual slides seems to be a great method to learn oral histology laboratory course. Based on the results of our study, we believe that the virtual microscopy with digitized virtual slides may gradually replace the real light microscopy with traditional glass slides for learning and teaching oral histology laboratory course. We foresee that virtual microscopy with digitized microscopic images will be extensively applied in many fields in the near future.

High-throughput whole-slide scanning to enable large-scale data repository building.

Digital pathology and artificial intelligence (AI) rely on digitization of patient material as a necessary first step. AI development benefits from large sample sizes and diverse cohorts, and therefore efforts to digitize glass slides must meet these needs in an efficient and cost-effective manner. Technical innovation in whole-slide imaging has enabled high-throughput slide scanning through the coordinated increase in scanner capacity, speed, and automation. Combining these hardware innovations with automated informatics approaches has enabled more efficient workflows and the opportunity to provide higher-quality imaging data using fewer personnel. Here we review several practical considerations for deploying high-throughput scanning and we present strategies to increase efficiency with a focus on quality. Finally, we review remaining challenges and issue a call to vendors to innovate in the areas of automation and quality control in order to make high-throughput scanning realizable to laboratories with limited resources. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Recent Advances in Pathology: the 2022 Annual Review Issue of The Journal of Pathology.

The 2022 Annual Review Issue of The Journal of Pathology, Recent Advances in Pathology, contains 15 invited reviews on research areas of growing importance in pathology. This year, the articles include those that focus on digital pathology, employing modern imaging techniques and software to enable improved diagnostic and research applications to study human diseases. This subject area includes the ability to identify specific genetic alterations through the morphological changes they induce, as well as integrating digital and computational pathology with 'omics technologies. Other reviews in this issue include an updated evaluation of mutational patterns (mutation signatures) in cancer, the applications of lineage tracing in human tissues, and single cell sequencing technologies to uncover tumour evolution and tumour heterogeneity. The tissue microenvironment is covered in reviews specifically dealing with proteolytic control of epidermal differentiation, cancer-associated fibroblasts, field cancerisation, and host factors that determine tumour immunity. All of the reviews contained in this issue are the work of invited experts selected to discuss the considerable recent progress in their respective fields and are freely available online (https://onlinelibrary.wiley.com/journal/10969896). © 2022 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Comparison of virtual microscopy and real microscopy for learning oral pathology laboratory course among dental students.

BACKGROUND/PURPOSE: Virtual microscopy has been used for teaching general and oral pathology laboratory course for more than 10 years. This study aimed to share the learning experience of an oral pathology laboratory course using either the virtual microscopy with digitalized virtual slides (virtual slide learning) or real microscopy using traditional glass slides (glass slide learning) among dental students. MATERIALS AND METHODS: Thirty-eight undergraduate dental students who took the compulsory course entitled "oral pathology" in the School of Dentistry, National Taiwan University were included in this study. The questionnaires were filled and final test was taken by these students after finishing the teaching of oral pathology laboratory course using either virtual or glass slide learning. The data were collected and analyzed statistically. RESULTS: Our results showed a significantly higher acceptance rate (all P-values < 0.001) and a significantly better histopathological diagnosis ability (P < 0.01) among dental students using the virtual slide learning than those using the glass slide learning for the oral pathology laboratory course. CONCLUSION: Virtual microscopy has many advantages over real microscopy in oral pathology laboratory course teaching. Based on the results of our study, we believe that the virtual microscopy with digitalized virtual slides may gradually replace the real microscopy with glass slides for the learning of oral pathology laboratory course. We foresee that the virtual microscopic images of the patients' specimens may be added in the patients' digital medical charts in addition to the images of computerized tomography, magnetic resonance imaging, and sonography in the future.

Digital mapping of Lewy bodies and neurites in alpha-synuclein stained large cerebral hemispheric sections from three patients with dementia with Lewy bodies showing psychotic manifestations: A pilot study.

BACKGROUND: Dementia with Lewy bodies (DLB) is one of the major neurodegenerative diseases and a clinical diagnosis is made based on the fourth consensus report on DLB. However, clinicopathological features of DLB are variable among cases. METHODS: We analyzed three autopsy-proven cases of DLB (patients 1-3). Their clinical features were variable in spite of their pathological commonality. The entire hemispheric sections were stained for phosphorylated alpha-synuclein (aS), digitized, and each aS positive lesion was mapped using a virtual slide system. RESULTS: The three patients were clinically diagnosed as having DLB. However, patient 1 exhibited amnesia and misrecognition, while patient 3 exhibited abnormal behavior in addition to dementia. Therefore, both patients 1 and 3 did not fulfill the clinical criteria of DLB, in contrast to patient 2. In spite of the clinical heterogeneity, Lewy pathology was similar in patients 1, 2, and 3. Additionally, patient 1 exhibited less frequent Lewy neurites of the amygdala and entorhinal cortex, and less frequent neurofibrillary tangles and senile plaque as compared to patient 2. On the other hand, the Lewy pathology of patient 3 extended far beyond those of patients 1 and 2, wherein the superior to middle frontal cortices, insular cortex, and lentiform nucleus were severely affected by Lewy pathology. CONCLUSIONS: Clinical features could be variable among autopsy-proven cases of DLB. Furthermore, we show that the accent of Lewy pathology differs according to the variability of the clinical symptoms. This method will provide a comprehensive strategy to analyze wide-spread aS lesions scattered throughout the entire cerebral hemisphere and help determine the corresponding pathological lesions responsible for the clinical variability of neurodegenerative disorders, including DLB.

Pathology without microscope: From a projection screen to a virtual slide.

We have witnessed successive stages since the Seventies in the advancements towards digital pathology. We agree with Dr Pallua et al on the tremendous changes that are taking place in pathology, all leading toward greater role of digitalization in the field of pathology, both in terms of consultation and teaching. In particular, distance teaching using digital pathology will grow into a mainstream mode of pathology teaching, something that has been reinforced by COVID-19.

Phosphomolybdic Acid Prevents Nonspecific Nuclear Staining by Picrosirius Red but Is Converted to Molybdenum Blue by Blue Light.

Picrosirius red (PSR) staining is generally used to evaluate liver fibrosis; however, PSR sometimes causes nonspecific nuclear staining. In this study, we evaluated the ability of phosphomolybdic acid (PMA) pretreatment to prevent nonspecific nuclear staining by PSR. In a manual evaluation of 27 non-tumor samples from patients with hepatocellular carcinoma, nonspecific nuclear staining was observed in 3.7% of PMA-treated specimens, compared with 85.2% of untreated specimens. Conversely, computer-assisted image analysis (CAIA) identified nonspecific nuclear staining in 0% of PMA-treated samples, vs 44.4% of untreated samples. Surprisingly, after mounting, PMA-treated specimens exhibited a blue tinge because of molybdenum blue (MB) production following sunlight exposure or virtual slide scanning. Using UV cut film, MB production induced by sunlight exposure was prevented; however, the film did not prevent MB production during virtual slide scanning. Moreover, only blue light-emitting diode exposure resulted in a blue tinge in PMA solution. Our data indicated that PMA pretreatment is effective for evaluating liver fibrosis using CAIA. Meanwhile, improvements in virtual slide scanning protocols would directly improve the quality of PMA-pretreated specimens subjected to CAIA.

Quantitative Analysis of Carbonic Anhydrase IX Uncovers Hypoxia-Related Functional Differences in Classical Hodgkin Lymphoma Subtypes.

Upregulation of carbonic anhydrase IX (CAIX) was found to be associated with unfavorable prognosis and resistance to treatment in a broad spectrum of malignancies, recently also in classical Hodgkin's lymphoma (cHL). As demonstrated, variable CAIX expression in a significant number of cHL cases was associated with poor treatment response. The current study focused on the quantification CAIX immunopositivity and its relative expression compared to the total CD30+ neoplastic pool using digital image analysis. One hundred and one lymph node samples featuring cHL histology were analyzed for both CD30 and CAIX by immunohistochemistry. Whole histological slides were scanned and immunopositivity was determined as the histoscore (H-score) using the DensitoQuant software module (3DHistech Kft., Budapest, Hungary). CAIX positivity was observed in the HRS-cells of 56/101 cases (55.44%) and frequently observed in the proximity of necrotic foci. CAIX H-scores were highly variable (range: 2.16-90.36, mean 18.7 ± 18.8). Individual CAIX values were independent of the much higher CD30 values (range 3.46-151.3, mean 52.37 ± 30.74). The CAIX/CD30 index proved to be the highest in the aggressive lymphocyte-depleted (LD) subtype (CAIX/CD30: 0.876). The CAIX expression and the CAIX/CD30 relative index can be precisely determined by image analysis, and values reflect the extent of a tumor mass undergoing hypoxic-stress-related adaptation in the most aggressive forms of cHL.

Pilot study on the correlation between dermoscopic patterns and fluorescence in situ hybridization findings using whole-slide digital imaging for acral volar melanocytic lesions.

Dermoscopic evaluation of acral volar skin is helpful in differentiating malignant melanomas (MM) from benign melanocytic nevi. However, histological diagnosis remains difficult because sufficient evidence of histopathological changes to establish a diagnosis of MM are not easily obtained. The aim of the present study was to evaluate the effective use of fluorescence in situ hybridization (FISH) in the diagnosis of acral volar melanocytic lesions, and to determine whether acral volar melanocytic lesions show characteristic molecular biological features of malignant melanoma via FISH. We classified acral volar melanocytic lesions showing junctional findings into three groups: (A) parallel ridge pattern (PRP) on dermoscopic examination with melanoma in situ; (B) PRP with insufficient melanocyte proliferation and atypia to diagnose malignant melanoma using hematoxylin-eosin staining; and (C) junctional nevi. We performed FISH analysis using the same tissue section that was used for hematoxylin-eosin staining. FISH positivity was seen in 80% (4/5) of the group A sections, and in 80% (4/5) of the group B sections. One case in group C was only 0.3% over the established criteria line (63.3% > 63% in RREB1). Our results suggest that FISH using whole-slide digital imaging may be useful in the diagnosis of early in situ MM when a typical PRP is observed in an acral volar skin lesion with non-diagnostic histopathology.

Brainstem tau pathology in Alzheimer's disease is characterized by increase of three repeat tau and independent of amyloid ß.

INTRODUCTION: Alzheimer-type neuropil threads (NTs) and neurofibrillary tangles (NFTs) are comprised of either 4 repeat (4R)-tau, 3 repeat (3R)-tau, or a mixture of both. In the hippocampus, the number of NFTs, and the proportion of 3R tau progressively increases. If this preferential accumulation of 3R tau also occurs in the brainstem, it may be fundamentally related to progression of Alzheimer pathology. METHODS: Midbrain and pontine sections of brainstems from 23 cases (Braak-NFT stages I/II: 8, III/IV: 8, and V/VI: 7) were double immunofluorolabeled for 4R and 3R tau. High-resolution (0.645 µm/pixel), in-focus snapshots were tiled to cover entire brain sections using a virtual slide system. Each lesion was classified by size (NT < 200 µm2 < NFT) and staining profile (3R/4R). In addition, the localization and quantity of amyloid ß (Aß) deposits were examined in adjacent sections for comparison with tau. RESULTS: The data sets obtained from approximately 286 gigabytes of image files consisted of 847,763 NTs and 7859 NFTs. The proportion of 3R tau-positive NTs and NFTs in the midbrain, and 3R tau-positive NTs in the pons gradually increased with advancing NFT stages, while the proportion of 3R tau-positive NFTs in the pons was already elevated at early stages. Aß deposits were absent at NFT stages I/II, and when present at later stages, their regional distribution was different from that of tau. These observations suggest that a progressive increase in the proportion of 3R tau occurs independently of Aß deposits. CONCLUSIONS: This is the first quantitative analysis of NFTs and NTs in the human brainstem. We demonstrate that the proportion of 3R tau in the brainstem neurofibrillary changes increases with disease progression. Because this phenomenon is shared between the brainstem and the hippocampus, this increase may be fundamental to the pathogenesis of Alzheimer disease.

Comparative analysis of fixation and embedding techniques for optimized histological preparation of zebrafish.

In recognition of the importance of zebrafish as a model organism for studying human disease, we have created zebrafish content for a web-based reference atlas of microanatomy for comparing histology and histopathology between model systems and with humans (http://bio-atlas.psu.edu). Fixation, decalcification, embedding, and sectioning of zebrafish were optimized to maximize section quality. A comparison of protocols involving six fixatives showed that 10% Neutral Buffered Formalin at 21°C for 24h yielded excellent results. Sectioning of juveniles and adults requires bone decalcification; EDTA at 0.35M produced effective decalcification in 21-day-old juveniles through adults (≥~3Months). To improve section plane consistency in sets of larvae, we have developed new array casting molds based on the outside contours of larvae derived from 3D microCT images. Tissue discontinuity in sections, a common barrier to creating quality sections of zebrafish, was minimized by processing and embedding the formalin-fixed zebrafish tissues in plasticized forms of paraffin wax, and by periodic hydration of the block surface in ice water between sets of sections. Optimal H&E (Hematoxylin and Eosin) staining was achieved through refinement of standard protocols. High quality slide scans produced from glass histology slides were digitally processed to maximize image quality, and experimental replicates posted as full slides as part of this publication. Modifications to tissue processing are still needed to eliminate the need for block surface hydration. The further addition of slide collections from other model systems and 3D tools for visualizing tissue architecture would greatly increase the utility of the digital atlas.

Proliferation of elastic fibres in idiopathic pulmonary fibrosis: a whole-slide image analysis and comparison with pleuroparenchymal fibroelastosis.

AIMS: We occasionally encounter patients with idiopathic pulmonary fibrosis (IPF) who have similar imaging patterns to those of pleuroparenchymal fibroelastosis (PPFE) in the upper lung fields but are not diagnosed as having PPFE clinically. The aim of this study is to identify the clinicopathological features and intrapulmonary distribution of elastic fibres and collagen fibres in these patients. METHODS AND RESULTS: We retrospectively reviewed the medical records of patients with a clinical diagnosis of IPF, and selected consecutive patients who underwent autopsy or pneumonectomy for lung transplantation. Patients with histologically confirmed PPFE were also reviewed for comparison. We quantified the collagen fibres and elastic fibres in each lobe as a percentage of the non-aerated lung area (collagen fibre score and elastic fibre score, respectively) in histological specimens by using whole-slide image analysis, and compared these scores between IPF and PPFE patients. In a total of 55 patients (IPF, 48; PPFE, 7), there were no significant differences in the collagen fibre scores between IPF and PPFE patients. The elastic fibre scores in the upper lobe in PPFE patients were significantly higher than those in IPF patients (23.5 versus 10.3, P = 0.005). However, it is of note that, in 12 of 48 IPF patients, the elastic fibre scores of the upper lobes were above the first quartile of those in PPFE patients. CONCLUSIONS: IPF occasionally shows intense elastosis in the upper lobes, and such cases are histologically indistinguishable from PPFE. There seem to be histologically borderline cases between PPFE and IPF.

PATMA: parser of archival tissue microarray.

Tissue microarrays are commonly used in modern pathology for cancer tissue evaluation, as it is a very potent technique. Tissue microarray slides are often scanned to perform computer-aided histopathological analysis of the tissue cores. For processing the image, splitting the whole virtual slide into images of individual cores is required. The only way to distinguish cores corresponding to specimens in the tissue microarray is through their arrangement. Unfortunately, distinguishing the correct order of cores is not a trivial task as they are not labelled directly on the slide. The main aim of this study was to create a procedure capable of automatically finding and extracting cores from archival images of the tissue microarrays. This software supports the work of scientists who want to perform further image processing on single cores. The proposed method is an efficient and fast procedure, working in fully automatic or semi-automatic mode. A total of 89% of punches were correctly extracted with automatic selection. With an addition of manual correction, it is possible to fully prepare the whole slide image for extraction in 2 min per tissue microarray. The proposed technique requires minimum skill and time to parse big array of cores from tissue microarray whole slide image into individual core images.

[Innovations in medical undergraduate pathology education: The Paris Descartes medicine faculty experience]. / Nouveaux outils et nouvelles méthodes pour l'enseignement de l'anatomie pathologique. Retour d'expérience à la faculté de médecine Paris Descartes.

At the Paris Descartes medicine faculty, we tested some newly developed tools to enhance the pedagogic value of the pathology teaching. In our faculty, this teaching is largely multidisciplinary and integrated in various teaching units; a large part is dedicated to practice works with thirteen 90min sessions. Virtual slides have been used for years in numerous medicine faculties; we successfully implemented this tool by adding contextual annotations, which facilitate students revising. We showed that rewarding students' assiduity enhanced their exam success. To do so, we now propose a short continuous assessment exam at the beginning of each practice session in the form of electronic multi-choice questions. Finally, we now propose a completely computerized final exam, on touchpads, that enhanced its docimologic value.

Comparison between Ki67 labeling index determined using image analysis software with virtual slide system and that determined visually in breast cancer.

BACKGROUND: In recent papers, Ki67 labeling index (LI) has been used to classify breast cancer patients into the low and high Ki67LI groups for comparison studies, which showed significant differences in many prognostic factors. It has not been clarified whether image analysis software can be used for calculating LI in breast cancer. In our study, we examined whether Ki67LI in breast cancer calculated using image analysis software correlates with that measured on the basis of visual. METHODS: Fifty patients were randomly selected among breast cancer patients who underwent surgical operation from March, 2010 to May, 2010 in our hospital without preoperative chemotherapy. In this study, for the virtual slide system (VSS: VS120-L100, Olympus, Tokyo, Japan), the high-resolution VSs of all the 50 patients were prepared as samples. The image analysis software use for calculating LI was Tissuemorph Digital Pathology (Tissuemorph DP: Visiopharm, Hoersholm, Denmark). The calculated LI was extracted from 3 to 5 views containing hot spots. The LI calculated using Tissuemorph DP was designed as LI/image/T. The digital image of 3 to 5 LI/image/T views was printed out, and on the digital photograph, we counted visually the number of Ki67-immunopositive cells in exactly the same area, and the percentage of Ki67-immunopositive cells was designed as LI/direct. Moreover, a pathologist's assistant (PA) determined the tumor area in the same specimen using VSS and calculated LI using Tissuemorph DP, which was designed as LI/image/PA. The chief pathologist (CP) similarly calculated LI which was designed as LI/image/CP. We evaluated the degree of agreement between different data sets "LI/image/T and LI/direct" and "LI/image/T, LI/image/CP, and LI/image/PA" by using interclass correlation coefficient (ICC). RESULTS: The average counts of cells were as follows: LI/direct, 3209.7 ± 1970.4 (SD); LI/image/T, 2601.6 ± 1697.1; LI/image/PA, 2886.5 ± 2027.5; LI/image/CP, 18805.5 ± 22293.4. The values of LI/direct and LI/image/T showed almost perfect agreement as showed by an ICC of 0.885 (95 % CI, 0.806-0.933; p < 0.001). The agreement among three investigators was almost perfect. The obtained ICC was 0.825 (95 % CI, 0.739-0.890; p < 0.001) among the data of LI/image/T, LI/image/CP and LI/image/PA. There were five cases that immunopositivity for Ki67 showed a more than 10 % disagreement between LI/direct and LI/image/T. CONCLUSION: The merits of calculating Ki67 LI using Tissuemorph DP are as follows. First, the staining intensity of the cells to be counted can be adjusted. Second, the portion of a tumor including "hot spots" for counting can be chosen. Third, many cancer cells can be counted more rapidly using Tissuemorph DP than by visual observation. However, it is important that pathologist should check and carry out the final decision of the data, when Ki67 LI using Tissuemorph DP is calculated.

[Improving practice in breast pathology: 34-months experience of the regional SENOPATH network and webinars as a tool for diagnosis of difficult lesions of the breast]. / Amélioration des pratiques en pathologie mammaire : bilan à 34 mois du groupe régional de relecture SENOPATH et télépathologie pour les lésions de diagnostic difficile.

Pathologists commonly face breast lesions that are difficult to diagnose. To reduce second opinion delay, erase geographical barrier and provide continuing education, we aimed to develop a telepathology-based regional network of pathologists. With the support of ONCOMIP network, we founded a peer-group named SENOPATH, composed of experienced breast pathologists practising in private laboratories, university hospitals or comprehensive cancer center in the region of Midi-Pyrénées in France. Submitted cases are digitalized at the University Hospital, stored in a shared space with a possible access via Internet prior to the SENOPATH sessions. The group meets monthly, via a synchronized webinar and multihead microscope session. A consensual diagnosis and final pathology report is issued for each case, and sent to the referring clinician via the patient medical file securely hosted by ONCOMIP. Between 2012 and 2014, 142 cases were reviewed, for either diagnostic 'routine' difficulty or rare histological type. The SENOPATH group, also regularly called by oncologists to solve difficult cases, has considerably improved the pathologist network in Southern France. Supported by the webinar tool, its educational impact is prominent, with a considerable progress in the region with regards to standardization of pathology processes, literature review and knowledge sharing.

Morphology in the Digital Age: Integrating High-Resolution Description of Structural Alterations With Phenotypes and Genotypes.

Conventional light microscopy has been used to characterize and classify renal diseases, evaluate histopathology in studies and trials, and educate renal pathologists and nephrologists. The advent of digital pathology, in which a glass slide can be scanned to create whole slide images (WSIs) for viewing and manipulating on a computer monitor, provides real and potential advantages compared with conventional light microscopy. Software tools such as annotation, morphometry, and image analysis can be applied to WSIs for studies or educational purposes, and the digital images are available globally to clinicians, pathologists, and investigators. New ways of assessing renal pathology with observational data collection may allow better morphologic correlations and integration with molecular and genetic signatures, refinements of classification schema, and understanding of disease pathogenesis. In multicenter studies, WSIs, which require additional quality assurance steps, provide efficiency by reducing slide shipping and consensus conference costs, and they allow slide viewing anytime and anywhere. Although validation studies for the routine diagnostic use of digital pathology still are needed, this is a powerful tool currently available for translational research, clinical trials, and education in renal pathology.

Support system for pathologists and researchers.

AIMS: In Japan, cancer is the most prevalent cause of death; the number of patients suffering from cancer is increasing. Hence, there is an increased burden on pathologists to make diagnoses. To reduce pathologists' burden, researchers have developed methods of auto-pathological diagnosis. However, virtual slides, which are created when glass slides are digitally scanned, saved in a unique format, and it is difficult for researchers to work on the virtual slides for developing their own image processing method. This paper presents the support system for pathologists and researchers who use auto-pathological diagnosis (P-SSD). Main purpose of P-SSD was to support both of pathologists and researchers. P-SSD consists of several sub-functions that make it easy not only for pathologists to screen pathological images, double-check their diagnoses, and reduce unimportant image data but also for researchers to develop and apply their original image-processing techniques to pathological images. METHODS: We originally developed P-SSD to support both pathologists and researchers developing auto-pathological diagnoses systems. Current version of P-SSD consists of five main functions as follows: (i) Loading virtual slides, (ii) making a supervised database, (iii) learning image features, (iv) detecting cancerous areas, (v) displaying results of detection. RESULTS: P-SSD reduces computer memory size random access memory utilization and the processing time required to divide the virtual slides into the smaller-size images compared with other similar software. The maximum observed reduction in computer memory size and reduction in processing time is 97% and 99.94%, respectively. CONCLUSIONS: Unlike other vendor-developed software, P-SSD has interoperability and is capable of handling virtual slides in several formats. Therefore, P-SSD can support both of pathologists and researchers, and has many potential applications in both pathological diagnosis and research area.

Applications and challenges of digital pathology and whole slide imaging.

Virtual microscopy is a method for digitizing images of tissue on glass slides and using a computer to view, navigate, change magnification, focus and mark areas of interest. Virtual microscope systems (also called digital pathology or whole slide imaging systems) offer several advantages for biological scientists who use slides as part of their general, pharmaceutical, biotechnology or clinical research. The systems usually are based on one of two methodologies: area scanning or line scanning. Virtual microscope systems enable automatic sample detection, virtual-Z acquisition and creation of focal maps. Virtual slides are layered with multiple resolutions at each location, including the highest resolution needed to allow more detailed review of specific regions of interest. Scans may be acquired at 2, 10, 20, 40, 60 and 100 × or a combination of magnifications to highlight important detail. Digital microscopy starts when a slide collection is put into an automated or manual scanning system. The original slides are archived, then a server allows users to review multilayer digital images of the captured slides either by a closed network or by the internet. One challenge for adopting the technology is the lack of a universally accepted file format for virtual slides. Additional challenges include maintaining focus in an uneven sample, detecting specimens accurately, maximizing color fidelity with optimal brightness and contrast, optimizing resolution and keeping the images artifact-free. There are several manufacturers in the field and each has not only its own approach to these issues, but also its own image analysis software, which provides many options for users to enhance the speed, quality and accuracy of their process through virtual microscopy. Virtual microscope systems are widely used and are trusted to provide high quality solutions for teleconsultation, education, quality control, archiving, veterinary medicine, research and other fields.