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Fixation on Histology: NSH Was Doing Distance Learning Before It Was Cool — Here’s Why It Still Works Written by: Connie Wildeman, MPA, Director of Education at NSH To Read the Full Blog, Click Here

Title: Histological Whole Slide Scanning Reproducibility Study   Authors: Hannah Benton, BSa, Tomoe Shiomi, MS, HTL(ASCP)CM, CT(IAC)CM, Fatma Farooqi, BSc, HTL(ASCP), Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC and Luis Chiriboga, PhD, HT(ASCP)QIHC Abstract: Whole slide imaging (WSI) is an increasingly versatile method for capturing and sharing high-resolution digital images of stained histological slides. These images can be used for a variety of applications, including clinical diagnosis, pathology review, and image analysis. While many whole slide scanners exist with varying features tailored to different use cases, a critical factor across all platforms is the accuracy and reproducibility of the scanned images. To investigate scan consistency over time, a control slide was prepared using a tissue microarray stained with Hematoxylin and Eosin (H&E). Ink dots were applied to the slide to define a consistent scanning region. The slide was scanned 77 times over six months using an Aperio AT2 whole slide scanner at 40x magnification.Image analysis was performed using HALO software by Indica Labs. Both the entire scan area and individual tissue punches were analyzed to assess total stained area and stain intensity, quantified by optical density (OD) for both hematoxylin and eosin. A linear regression model was applied to data from all individual punches and the full scan region. Additionally, a two-way ANOVA was conducted to compare OD values of hematoxylin and eosin between the first 10 scans and the last 10 scans. Key findings were that hematoxylin showed a statistically significant decline in both stained areas and OD over time, while eosin demonstrated a statistically significant increase in stained area, but a decrease in OD. These results suggest potential degradation of staining quality or imaging consistency over time. Possible contributing factors include slide bleaching, light source variability, annotation region size, or other imaging conditions. These will be the focus of future investigations to better understand and control variability in longitudinal slide scanning studies.

Title: Employing Multi-Tissue Controls to Enhance Kidney Biopsy Protocol Education in a Program in Histotechnology Student Lab  Authors: Hyder Aljanabi, Damon Bendolph, Gabriella Casas, Yosan Embrafrash, Sara Hassan, Anastasja Kraft, Stephan Lloyd-Brown , Nida Mubeen, Minh Nguyen, Xena Orosco, Nicole Rivera, Moriam Sissoho, Tan Tang , Kaleena Ramirez, Toysha Mayer, Mark Bailey Abstract: In a Program in Histotechnology student laboratory, establishing a representative and clinical teaching laboratory environment is essential for preparing students to manage the complexities of diagnostic tissue processing. The objective of the project was to simulate real-world clinical procedures by integrating multi-tissue controls into student education competencies for kidney biopsy staining protocols. Students participated in the investigation, each receiving four pieces of formalin-fixed, paraffin-embedded (FFPE) tissue: kidney, liver, gastrointestinal tract (GI), and tonsil. The tissues served as controls to validate staining techniques commonly used in renal pathology. Students prepared tissue sections using a rotary microtome, sectioning tissue at four microns. In total, forty slides were prepared, with eighteen slides manually stained using specific histochemical methods. Stains included hematoxylin and eosin (H&E), periodic acid-Schiff (PAS), periodic acid methenamine silver (PAMS), and the Gomori Trichrome technique. The results yielded identifiable cellular and structural features critical for diagnostic interpretation. A slide review was conducted, and acceptable representative slides were selected for digital imaging. In addition, the results demonstrated the four tissue types which may be approved to use as controls, due to the consistency of demonstrating staining characteristics and features required for evaluating kidney biopsy protocols. Upon technical validation, the use of multi-tissue controls contributed to educational and operational outcomes. Students gained quality assurance experience, and the experience reinforced special stain and laboratory operations competencies, teaching students how to conserve reagent use, and to reduce time and expense. Furthermore, the protocol introduced the application of digital pathology and quality assurance in a real-world lab setting. Our investigation supports the integration of multi-tissue controls in histotechnology education as a valuable tool for enhancing both learning and laboratory efficiency. Future studies are recommended to include additional tissue types, stains, and immunohistochemical markers, to further advance and expand histotechnology educational competencies.

Title: Comparative assessment of routine H&E and Mason’s Trichrome Stain to Differentiate Normal from Infected Urinary Bladders in the Göttingen Minipig   Authors: Stephanie D. Rivera, MS, HT(ASCP); Anthony Romanello, BS; Ronnie Chamanza, BVSc, MSc, FRC Path, MRCVS, FIATP, Preclinical Sciences & Translational Safety, Johnson & Johnson Innovative Medicine, Spring House, Pa Abstract: The objective of this experiment was to evaluate the difference between normal and infected Göttingen minipig urinary bladder and determine the difficulty of microscopically evaluating infectious urinary bladder. The focus was on the histology of the tissue using routine Hematoxylin and Eosin(H&E) staining and the Masson Trichrome (MT) special stain to demonstrate the morphology of normal and infectious urinary bladder. To improve on the translation between preclinical and clinical studies, the Göttingen minipig model is appropriate to use for research for diseased human bladder treatment because the minipig anatomy and organ system are similar to humans. The procedure was designed to evaluate normal uninfected urinary bladder and bacteria infected (E. coli) urinary bladder by evaluating morphology/cellular changes associated with the resultant inflammatory response in the urinary bladder of the Ellegaard Gottingen Minipig. The H&E-stained urinary bladder tissue section and a Masson Trichrome stained urinary bladder section were used to evaluate control ‘normal’ tissue vs infected tissue cellular differences. Optimal microscopic evaluation requires that the urinary bladders are properly fixed and processed. By using proper fixation and diligent histologic practices, all components of the urinary bladder are captured for proper histologic evaluation.

Title: Evaluating the Reverse Slide Embedding Method vs. Heat Extractor Embedding in the Mohs Laboratory: A Comparative Quality Review of 100 Cases Authors: Tashsa Cromedy, Heather Frye, Ochsner MD Anderson Cancer Center, St. Tammany Cancer Center A Campus of Ochsner Medical Center Abstract:  Overview Accurate tissue embedding is critical in Mohs micrographic surgery for complete margin assessment. This study evaluates the efficacy of a reverse slide embedding method compared to the conventional heat extractor technique. The goal was to determine which method yields fewer artifacts or discrepancies that may compromise histologic interpretation and margin assessment. Methods A total of 100 Mohs cases were retrospectively reviewed in a controlled laboratory setting. Two embedding techniques were compared: Reverse Slide Method: 50 cases were embedded by placing the tissue on a chilled slide before embedding, ensuring orientation preservation and minimizing heat exposure. Heat Extractor Method: 50 cases were embedded using the traditional heat extractor to flatten and orient tissue in the embedding medium. All slides were reviewed by a Mohs surgeon for processing artifacts, orientation challenges, and histologic discrepancies. Validation The Mohs surgeon identified a total of 17 artifact inconsistencies or discrepancies across all cases: 13 instances were associated with the heat extractor method. 4 instances occurred with the reverse slide method. These findings suggest that the reverse slide method may reduce artifacts and improve embedding accuracy compared to the heat extractor, offering potential benefits for tissue integrity and diagnostic confidence in the Mohs laboratory. Conclusion The reverse slide embedding method demonstrated a significant reduction in embedding-related artifacts compared to the heat extractor technique. These findings support its use in the Mohs laboratory to enhance tissue quality, reduce the risk of diagnostic errors, and improve patient outcomes. Further studies with larger sample sizes and multi-lab validations are recommended to confirm these results.  

Title: High-resolution histological preparation of Araneomorphae and Mygalomorphae chelicerae using a modified petrographic technique Authors: Damien Laudier, HTL(ASCP)QIHC, Laudier Histology Abstract: Producing quality histological preparations of spider chelicerae with articulated fangs and cheliceral teeth is exceptionally challenging, if not impossible, using conventional histology techniques. Typically, these structures are examined with topographic or radiographic imaging methods, such as scanning electron microcopy (SEM) and micro computed tomography (mIcro-CT). While both are very useful tools for morphological analysis, they’re not capable of revealing the fine tissue structure and cellular details, that a histological section viewed under light microscopy can provide. This study describes a modified petrographic/hard tissue histology technique to prepare high-resolution histology sections, for qualitative and quantitative assessment of both cheliceral soft tissue and fang microstructure.

Title: Enhanced Collagen Detection in Liver Fibrosis: A Comparative Study of Picrosirius Red Staining With and Without Bouin’s Pretreatment   Authors: Nate Rampy, BS, Amber Moser, BS, HTL(ASCP)cm, Hannah Benton, BS, Brad Bolon, DVM, MS, PhD, DACVP, DABT and Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC, Premier Laboratory, LLC, Longmont, Colorado; GEMpath, Inc., Longmont, Colorado Abstract: The use of Bouin’s solution as a post-fixation treatment, rather than a primary fixative, remains largely unexplored in Picrosirius Red (PSR) procedures for collagen detection. In this study, we compared the effectiveness of PSR staining in liver samples from mouse, rat, and human with and without Bouin’s solution as a pretreatment step. Liver sections were fixed in 10% neutral buffered formalin, processed and embedded in paraffin before being sectioned at 4 microns and stained with PSR. Bouin’s was applied prior to staining for 60 minutes at 70º C, not as a fixative, but as a mordant to enhance dye-tissue interactions. Stained slides were scanned at 20x with an Aperio AT2. Visual assessment and image analysis in bright field microscopy demonstrated that the slides pretreated with Bouin’s had significantly improved collagen differentiation, with enhanced contrast. By comparison, slides stained without the Bouin’s pretreatment showed weaker and less distinct collagen staining. Our findings suggest that Bouin’s pretreatment significantly improves collagen staining contrast and differentiation. The use of Bouin’s pretreatment may serve as a valuable revision to the standard histology protocol for PSR fibrosis evaluation as well as general collagen visualization.

Title: Wheat Germ Agglutinin (WGA) Staining Optimized for Image Analysis of Muscle Tissue Morphometry  Authors: Cheru, R. and Wolf, J.C., Experimental Pathology Laboratories, Inc., Sterling, Virginia Abstract: Wheat Germ Agglutinin (WGA) is a plant-derived lectin and fluorescent stain that binds to N-acetylglucosamine and sialic acid residues in tissues, making it a valuable histochemical tool for visualizing cell membranes and components of the extracellular matrix. In muscle tissue, WGA staining allows clear delineation of the laminin-labeled basal membrane outlining each myofiber, distinguishing it from the residual autofluorescence of the myofiber sarcoplasm. To support digital pathology applications, a WGA staining protocol was optimized for compatibility with image-based quantitative analysis. Formalin-fixed, paraffin-embedded muscle sections were stained with fluorescently labeled WGA, counterstained with DAPI for nuclear visualization, and mounted with antifade medium to preserve fluorescence. Image analysis of WGA-stained skeletal muscle was successfully performed by a pathologist using Image-Pro® Plus software, employing macros to assess myofiber size and count.

Poster Title: CelLockTM and Axlab: A mutual symbiotic relationship resulting in increased patient care quality. Authors: Clifford M Chapman ( Medi-Sci Consultants), Karla Escobar (Axlab -US), Timm Piper (Axlab –US) Abstract: An ever increasing number of tiny specimens are received in histology laboratories every day. Fine needle aspirates, needle biopsies, gastrointestinal and skin biopsies, along with research specimens such as organelles: all pose challenges in receiving, processing and preparing stained slides from such specimens. Axlab is a Danish based company founded in 1993 which specializes in finding novel solutions for pathology laboratories. Axlab recently announced the release of their AS-410M automated sectioning equipment. Receiving FDA clearance in 2024, this equipment has been successfully implemented worldwide, resulting in enhanced section quality and increased workflow efficiency. CelLockTM is an innovative standardized method for collecting individual cells and small tissue fragments for subsequent routine, immunohistochemical and molecular pathology diagnostic and investigative techniques. The CelLock method, which utilizes a novel product CelLGelTM , results in the collection and retention of 99.9% of the original specimen within a paraffin embedded cell-block. In addition, the specimen is precisely located within the paraffin block, close to the surface, and is identified by a marker to cue when sections should be taken. Axlab is currently investigating the use of CelLock and CelLGel in the preparation of cell blocks which can be precisely sectioned on their automated sectioning equipment. The initial results are presented in this poster.

Poster Title: The High Cost of Understaffing: A Case Study in Surgical Pathology Consequences   Authors: Emily Nangano, MS, PA(ASCP)cm; Gillian Bass; Rob Terranova Abstract: Laboratories are the diagnostic backbone of healthcare, yet staffing decisions are often driven by budget constraints rather than operational needs. This case study examines the real-world consequences of delayed staffing action within the anatomic pathology department at a large academic medical center. Faced with a predicted shortfall in grossing coverage due to reduced resident support and unchanged PA staffing levels, institutional leadership opted against proactive hiring. As a result, grossing FTEs fell from 6.5 to 3.5, and histology staffing experienced a drop to 3 technicians from the usual 9 due to attrition and burnout. This staffing collapse led to turnaround time delays of up to 6–8 weeks and forced the lab to outsource specimen processing. Over the following seven months, the institution spent nearly $4 million on reference lab services. Staff morale declined sharply, clinician trust eroded, and senior PAs and histotechs resigned. Even after additional staff were hired, it took more than a year to stabilize operations. This poster presents supporting data, including FTE changes, outsourcing costs, and turnaround time impacts. It also explores how temporary, qualified locum tenens staffing solutions—such as Pathologists’ Assistants and histotechnologists, and cytologists—can help bridge coverage gaps and prevent costly disruptions. Ultimately, this case underscores the critical importance of timely, proactive staffing strategies. The hidden costs of under-resourcing the laboratory go beyond dollars—they affect staff well-being, institutional reputation, and patient care outcomes.