Search

Antigen-bearing proteins become progressively unavailable to immunodetection after prolonged storage of routine sections, exposed to a variety of agents, such as moisture, oxygen, and temperature. By proteomic analysis, the antigens are retained in the sections and definitely in the tissue block, pointing to fixation-independent, storage time–dependent protein modifications. Based on previous experience, we hypothesized that a combined exposure to a reducing agent and to chemicals favoring protein conformation changes would reverse the masking in aged sections. Disaccharides, lactose and sucrose, and a surfactant, added to a standard antigen retrieval buffer, reverse the negative changes in aged sections. Furthermore, they provide enhanced access to antigens in freshly cut sections, but not universally, revealing additional factors, besides heat and calcium chelation, required for antigen retrieval of individual proteins

Whole-slide images (WSI) have acquired a stable place in diagnostic histopathology and immunohistochemistry. Immunofluorescence (IF) techniques hold a limited and selective role in diagnostics (eg, renal and cutaneous pathology) and so far remain excluded from the digital pathology evolution, with notable exceptions, such as quantitative immunopathology. We explored the ability of a commercial fluorescent slide scanner to provide 4-color IF WSI from routinely processed tissues. With minor modifications and a careful match between filters and fluorochromes, we show that 4-color IF WSI can be obtained from routine material with negligible autofluorescence, good sensitivity, and diagnostic power.

Aims The aim of this study was to investigate whether the shelf-life of diagnostic antibodies is longer than the expiry date on the label. Methods and results Four independent laboratories tested a small number of diagnostic antibodies kept at +4°C for 12–26 years, and found them to work perfectly on routine histology sections. Conclusions Diagnostic antibodies may have a workable half-life in excess of 10 years, and the emphasis on performance should shift to the preservation of antigenic targets in the tissue.

Background The International Society of Nephrology/Renal Pathology Society (ISN/RPS) classification represents the gold standard for the histological evaluation of Systemic Lupus Erythematosus (SLE) nephritis. A repeat biopsy (RB) might be an important tool to provide information on long-term renal outcomes and optimal therapy. Aims of this study were to evaluate the use of the ISN/RPS classification and the role of RB in routine clinical practice. Methods A total number of 142 patients with SLE nephritis and with adequate reference and RB samples were included in this multicentre retrospective study. A meticulous histological examination was centrally performed on first and RB and compared with clinical variables and follow-up data. Results Morphological features of the ISN/RPS classification: at first and RB, significant differences were observed between segmental classes (III, IV-S) and Class IV-G in mesangial proliferation, wire loops and tuft necrosis. Clinical features and ISN/RPS classification: the correlation between serum creatinine, proteinuria, blood pressure levels and histological classes at first and RB demonstrated more severe renal disease in Class IV-G, both at first and RB. Agreement between ISN/RPS classification at first and RB: 40.8% of patients changed the histological class. Fifty per cent of Class II (mild mesangial form) were reclassified as Class IV-G at RB, whereas 18.9% of Class IV-G were reclassified as Class II. The transition among segmental (III/IV-S) and mesangial forms (II/IV-G) was extremely rare. The comparison between the clinical parameters at the final follow-up and the ISN/RPS classification confirmed that the trend of serum creatinine and proteinuria between the different classes was better described at the RB (higher in Class IV-G) than on the first biopsy. Conclusions The histopathological data suggest that morphological differences between segmental and global forms do exist, possibly due to different pathogenetic mechanisms. An RB strategy could provide additional information on long-term renal outcomes. A strategy of protocol biopsies could be useful in perspective future trials to better understand the therapeutic response and the natural history of this disease.

The international guidelines emphasize the crucial role of renal biopsy in the diagnosis of Anti-Neutrophil Cytoplasmic Autoantibody (ANCA)- associated glomerulonephritis (Microscopic Polyangiitis, Granulomatosis with Polyangiitis and Eosinophilic Granulomatosis with Polyangiitis). 1) According to the recent 2012 Chapel Hill Consensus Conference, ANCA- associated vasculitides are classified in the group of small vessel vasculitis. 2) Pauci-immune necrotizing crescentic glomerulonephritis is the morphological hallmark of ANCA-associated vasculitis. The lesion can vary widely in severity and extent of the damage, from segmental tuft necrosis to massive circumferential crescents and frequent peri-glomerular granulomatous reaction. 3) The ANCA test is highly specific for these types of autoimmune vasculitides but renal biopsy still remains the gold standard for final diagnosis, prognostic evaluation and therapeutic intervention, although discrepancy between clinical and morphological features is frequently found. 4) Crescentic damage of the glomerular tuft is characterized by monocyte.- macrophage accumulation through Vascular Cell Adhesion Molecule-1 (VCAM-1) activation. Activated macrophages are considered to have a key role in the chronic progression of renal damage due to their ability to produce substances directly involved in matrix remodelling (Tumor Growth Factor b). 5) Diffuse and marked interstitial infiltration of T, B lymphocytes and monocyte- macrophages is another frequent morphological feature with aspects of tubulitis recently being considered important markers for a worse prognosis. 6) Unfortunately, repeat biopsies are infrequently performed in these disorders and long-term renal changes have remained largely unidentified. 7) Despite the formulation of standardized scoring for renal biopsies, definitive histopathologic classification is still lacking. The European Vasculitis Study (EUVAS) group has proposed a classification system based on glomerular pathology as assessed by light microscopy which is, in turn, divided into four categories: focal, crescentic, sclerotic and mixed. A preliminary correlation with clinical features in 100 cases of ANCA-associated vasculitis has demonstrated that renal biopsy categories were the only independent predictors for the estimated glomerular flow rate (eGFR). The international study currently way is being evaluated by EUVAS for possible confirmation of the classification.

Context: In 2013 the high throughput technology known as Tissue Micro Array (TMA) will be fifteen years old. Its elements (design, construction and analysis) are intuitive and the core histopathology technique is unsophisticated, which may be a reason why has eluded a rigorous scientific scrutiny. The source of errors, particularly in specimen identification and how to control for it is unreported. Formal validation of the accuracy of segmenting (also known as de-arraying) hundreds of samples, pairing with the sample data is lacking. Aims: We wanted to address these issues in order to bring the technique to recognized standards of quality in TMA use for research, diagnostics and industrial purposes. Results: We systematically addressed the sources of error and used barcode-driven data input throughout the whole process including matching the design with a TMA virtual image and segmenting that image back to individual cases, together with the associated data. In addition we demonstrate on mathematical grounds that a TMA design, when superimposed onto the corresponding whole slide image, validates on each and every sample the correspondence between the image and patient′s data. Conclusions: High throughput use of the TMA technology is a safe and efficient method for research, diagnosis and industrial use if all sources of errors are identified and addressed.

Cancer derives from a cell clone that has accumulated genetic and epigenetic changes that influence its phenotype and finally enable it to escape from the normal controls of proliferation. A recent paper shows that, in myc-induced tumours, the inactivation of this oncogene produces the regression of the tumours and the differentiation of the tumour cells into mature osteocytes.1 In addition, a further reactivation of myc in these cells does not restore the malignant phenotype but induces apoptosis. This discovery could lead to an innovative therapeutic strategy. BioEssays 25:104–107, 2003. © 2003 Wiley Periodicals, Inc.