Neutral-buffered Formalin Research Articles

Clinico-histopathological and phylogenetic analysis of protozoan epibiont Epistylis wuhanensis associated with crustacean parasite Lernaea cyprinacea from ornamental fish in Iran

Parasitic infestations are one of the most concerning problems limiting ornamental fish farming. In addition to the direct economic losses due to the major mortalities, parasites may significantly negatively impact the body shape, coloration, growth rate, and health condition of the fish. The results of the study highlight the importance of comprehensive parasitological analysis in the diagnosis and treatment of parasitic infections in ornamental fish farms. The presence of multiple parasites in the affected fish emphasizes the need for effective biosecurity measures, such as extending the quarantine period for newly imported fish, closely monitoring fish populations, and implementing isolation units to prevent the spread of infections. By implementing these preventative measures, ornamental fish farmers can reduce the risk of parasitic infections and ensure the health and well-being of their fish populations. This, in turn, can lead to increased profitability and sustainability for their business. Overall, the current study aimed to conduct a clinical, histopathological, and phylogenetic analysis of the epibiont ciliated protozoan Epistylis wuhanensis and the copepod crustacean Lernaea cyprinacea in a freshwater ornamental fish farm in Iran. Furthermore, it provides valuable insights into the prevalence and impact of parasitic infections in ornamental fish farms and underscores the need for continued research and the development of effective preventative measures to address this issue. A total of 60 symptomatic freshwater ornamental fish, including 30 guppy (Poecilia reticulata) and 30 sailfin molly (Poecilia latipinna), were packed in polyethylene bags filled with oxygenated pond water and transported to the Ornamental Fish Clinic, Faculty of Veterinary Medicine, University of Tehran, for parasitological analysis. Following the clinical examination, histopathological analysis was performed on 10% NBF (neutral buffered formalin)-fixed samples from affected tissues, including the skin, skeletal muscle, and liver, to identify any pathological changes associated with the parasitic infections. Furthermore, the DNA was extracted from the 99% ethanol-fixed samples using a commercial DNA extraction tissue kit (SinaPure DNA, Iran), and PCR was performed using Peri18S-F1 (5′-ACC TGG TTG ATC CTG CCA GT-3′) and Peri18S-R1 (5′-TGC AGG TTC ACC TAC GGA AA-3′) (first reaction), and Peri18S-F2 (5′-CCG CGG TAA TTC CAG CTC-3′) and Peri18S-R2 (5′-GAT CCC CTA ACT TTC GTT CTT GA-3′) (second round) primers for the identified parasites. Finally, the PCR products were sequenced using Sanger dideoxy sequencing methods, and the resulting sequences were compared to sequences in the BLAST search program to provide a comprehensive picture of the current parasite-based disorder. The crustacean L. cyprinacea and the epibiont sessilid E. wuhanensis were identified in the examined ornamental guppy (6/30) and sailfin molly (6/30), with an overall parasitic prevalence of 20.00% (12/60). Ciliates were found in all tissue lesions but not in fish without lesions. A great number of the ciliated protozoan E. wuhanensis were found attached to the integumentary area of L. cyprinacea. Microscopically, oval to round granulomatous lesions were observed in cutaneous and skeletal muscles. Lymphoplasmacytic dermatitis and myositis were also observed. The crustacean L. cyprinacea serves as a mechanical vector for E. wuhanensis infection and spreads the disease in ornamental fish farming operations. For the first time in Iran, we successfully presented diagnostic morphological and molecular data on sessilids isolated from L. cyprinacea. Based on the findings of the current study, such parasitic infections may cause significant economic losses following invasion of the integument area of fish, eventually leading to death, if treatment is neglected or inadequate. Furthermore, the findings of the analysis were used to develop effective diagnostic approaches for the affected fish, as well as recommendations for improved health conditions to prevent future outbreaks of parasitic infections. However, further research is needed to determine the precise mechanisms of crustacean attachment and host-crustacean-peritrich protozoan interactions. Furthermore, the direct and indirect effects of various environmental factors on the emergence and spread of the current disease should be considered.

Effectiveness of neutral honey as a tissue fixative in histopathology

Background: In routine histopathology, 10% neutral buffered formalin (NBF) is the choice fixative. However, formalin is a human carcinogen, so there is a necessity for a safer alternative. To the best of our knowledge, neutral honey, where the pH is around seven, not natural or artificial honey, has not been tested to fix histological samples. This study determined the effectiveness of neutral buffered honey and other types of fixatives to fix histological tissues. Methods: The study was conducted between July 2019 and August 2020 at Sultan Qaboos University, Oman. Sections from three rat livers, kidneys, and stomach tissues were fixed with 10% NBF, neutral buffered Sumer honey (dwarf honey bee), neutral buffered date honey, formalin, Sumer honey, date honey, alcoholic formalin, alcoholic Sumer honey, and alcoholic date honey for 24 hours. Samples were stained with hematoxylin and eosin (H&E), special stains, and vimentin methods. Three expert biomedical scientists then evaluated the fixed and stained samples for the quality of all sections. The fixation ability of the different honey solutions was then compared to 10% NBF and the utility was determined using nuclear and cytoplasmic criteria, specificity, and intensity. Results: H&E showed adequate nuclear staining in all groups compared to 10% NBF. The specificity and intensity of all groups for the Periodic acid–Schiff method were identical to 10% NBF except for Sumer honey and alcoholic date honey. Vimentin showed comparable findings with 10% NBF as there were no significant differences. Conclusions: The findings of this study encourage the use of honey, including neutral, as a possible safe substitute fixative for formalin, however, further experiments on larger specimens should be conducted.

Study of significance of p53 expression in primary ovarian tumour

The purpose of the study was to understand the molecular changes that happen in human cancer, P53 gene mutation. Here, with respect to P53 immunohistochemical staining were taken into consideration as the surrogate for the mutational analysis. This was used for the diagnosis of ovarian cancer. The main aim of the study was to evaluate the immunohistochemical frequency of the P53 and to correlate its expression with the prognosis. In the current study 107 cases associated with ovarian cancer were studied. The samples received were hysterectomy as well as unilateral and bilateral salpingo-oophorectomy specimens, which were fixed in 10% neutral buffer formalin and processed routinely. 68.7% of the patients showed P53 malignant ovarian tumours. In the benign, it was considered as negative or in the position of borderline tumour. No statistically significant correlation was shown between the age and P53 expression. There was statistically significant difference between borderline, benign and the malignant tumors. It was also assessed that there was a statistically significant difference between P53 expression and serous carcinoma in the current study. The present study concludes that P53 tumor gene was found to be mutated in more than the 50% of the human cancer and was expressed as the aberrant form of protein. The study also concludes that in borderline tumor, P53 rarely mutates and is results in poor prognosis.

Histopathologic survey of free-living populations of 2 species of freshwater mussels in Indiana.

Freshwater mussels are one of the most endangered groups of animals in Indiana, with nearly half of the native species either extirpated or listed as "state endangered" or of "special concern." Nationally, numerous freshwater mussel species are considered threatened. Freshwater mussel diseases are not well understood and few published accounts of freshwater mussel diseases with detailed histological descriptions exist. Mass mortality events within mussel populations are increasingly recognized, often with undetermined etiology. Our objective was to determine baseline histopathology in free-living populations of freshwater mussels. One-hundred twenty individual mussels representing 2 species-plain pocketbook (Lampsilis cardium) and fatmucket (Lampsilis siliquoidea)-were collected from 3 different locations within the Wildcat Creek watershed in central Indiana during June and July 2019. A cross-section through the visceral mass was obtained and immersed in 10% neutral-buffered formalin, with routine processing and hematoxylin and eosin staining. Branchial acariasis occurred in 43/60 fatmuckets and 22/60 plain pocketbooks. Infection with a bucephalid trematode was recognized in 18/60 fatmuckets, while infection of the gonadal duct with an unidentified trematode species was identified in 4/60 fatmuckets and 18/59 plain pocketbooks. Additional changes associated with unidentified trematodes, bacteria, fungi or oomycetes, and ciliates were observed. Other miscellaneous changes included mineralization, neuronal lipofuscinosis, and gonadal atrophy/atresia. A range of histological changes were observed. These changes likely represented background lesions: incidental findings, spontaneous infectious or endosymbiotic conditions, or normal physiological changes that routinely occur in free-living wild populations. Awareness of baseline lesions should inform future diagnostic investigations of mussel mortality events.

Morpho-histological Study of the Nictitans Gland and Cartilage in Dogs and Cats in Sulaimani Province

This comparative study aims to assess the differences in the third eyelid gland and cartilage morphology and histology between dogs (Canis lupus familiaris) and cats (Felis catus). Twenty nictitans membranes were collected from 20 adult dogs and 20 adult cats. The nictitans membranes were dissected to examine the structure, shape, and positions of the nictitans gland and cartilage. Histologically, the samples were fixed in 10% neutral buffer formalin and then prepared for H&E stains. Generally, in both species, the cartilage consists of a long, slender body, and pigmented large dorsal and small ventral wings. The cartilage had a curving shape that resembled the eyeball's curvature and had two surfaces; palpebral and corneal or bulbar surface. At the basal end of the corneal surface, there are several lymph nodes aggregates. In the dogs, the nictitans cartilage was longer and slenderer than in the cats. The body of the cartilage was wrapped by a spherical and light pink nictitans gland in the dogs while the cats had oval nictitans gland. Microscopically, the nictitans gland in the dogs was composed of a compound tubuloacinar gland whereas; in cats, it was composed of a compound acinar gland. There was no difference histologically in the intralobular and the interlobar septa between species, the isogenous group of chondrocytes was higher in cats Vs. the dogs that showed few in number. In conclusion, the nictitans gland and cartilage were varied in their anatomical and histological structure. This should help researchers make preparations to prevent negative impacts and could help them investigate changes made more pronounced by disease conditions.

Diagnostic application of formalin fixed archived tissues for detection of foot-and-mouth disease

Early and definitive disease diagnosis is critical for effective disease control. 50% buffered glycerine is commonly used viral transport medium, which is not always available and required cold chain. Tissues samples archived in 10% neutral buffered formalin (NBF) can preserve nucleic acid that can be used in molecular studies and disease diagnosis. The present study's goal was to detect the foot-and-mouth disease (FMD) viral genome in formalin-fixed archived tissue which may avoid cold chain during transportation. This study used FMD suspected samples preserved in 10% neutral buffered formalin from 0 to 730 days post fixation (DPF). All archived tissues were positive for FMD viral genome by multiplex RT-PCR and RT-qPCR up to 30 DPF, whereas archived epithelium tissues and thigh muscle were positive for FMD vial genome up to 120 DPF. FMD viral genome was detected in cardiac muscle up to 60 DPF and 120 DPF, respectively. The findings suggest that 10% neutral buffered formalin could be used for sample preservation and transportation for timely and accurate FMD diagnosis. More samples need to be tested before implementing the use of 10% neutral buffered formalin as a preservative and transportation medium. The technique may add value in ensuring biosafety measures for creation during disease free zone as well.

A New Medium (HistoCold) for Surgical Specimens Preserving to Improve the Preanalytic Issues in Histopathological Samples Handling: Morphologic and Antigenic Analysis.

Introduction: The onset of precision medicine has led to the integration of traditional morphologic tissues evaluation with biochemical and molecular data for a more appropriate pathological diagnosis. The preanalytic phase and, particularly, timing of cold ischemia are crucial to guarantee high-quality biorepositories of formalin-fixed paraffin-embedded (FFPE) tissues for patients' needs and scientific research. However, delayed fixation using the gold-standard and carcinogenic fixative neutral-buffered formalin (NBF) can be a significant limitation to diagnosis and biopathological characterization. HistoCold (patented; Bio-Optica Milano S.p.A., Milano, Italy) is a nontoxic, stable, and refrigerated preservative solution for tissue handling. This study examined HistoCold's potential role in improving the preanalytic phase of the pathological diagnostic process. Materials and Methods: Breast, lung, or colorectal cancers (20, 25, and 10 cases, respectively) that were to be surgically resected were recruited between 2019 and 2021. Once specimens were surgically removed, three residual samples for each patient were first promptly immersed into HistoCold for 24, 48, and 72 hours and then FFPE. These were compared with routine specimens regarding morphologic features (hematoxylin and eosin) and tissue antigenicity (immunohistochemical stains). Results: Good concordance regarding both the morphologic characteristics of the neoplasms and their proteins expression between the routine and HistoCold handled tissues were found. The tissue handling with the solution never affected the histopathological diagnosis. Conclusions: The use of HistoCold for samples transporting is easy, allows for improving the management of cold ischemia time, and monitoring the fixation times in NBF, resulting in good quality tissue blocks for biobanking. Moreover, it could be a candidate to eliminate formalin from operating theaters. HistoCold looks very promising for the preanalytic phase of human tissues handling in the era of precision medicine, to provide the best service to patients, and to scientific research.

Standardization of Grocott's methenamine (hexamine) silver method for glycogen demonstration in liver tissue.

Demonstration of glycogen can be done in different lesions and is considered diagnostically significant, mainly in some tumors. Glycogen staining is affected by the type of fixative, the temperature of fixation, and the staining technique.Grocott's methenamine (hexamine) silver technique quality was assessed after four different types of fixatives at two different temperatures [Bouin's solution, 10% neutral buffered formalin (NBF), 80% alcohol, and Rossman's solution at room temperature (RT) and 4°C, for 24h]. These variables were studied to optimize this technique for glycogen demonstration. Archived paraffin blocks were used in this study. They were prepared from one rabbit's liver, and 32 paraffin sections were prepared and stained with Grocott's methenamine (hexamine) silver technique. Eightypercent alcohol provided the highest staining quality scores at both RT and 4°C in comparison with the other fixatives. We concluded that 80% alcohol at 4°C seems to be the fixative of choice for glycogen with the Grocott's methenamine (hexamine) silver technique at the level of this study.

Simple Universal Whole-Organ Resin-Embedding Protocol for Display of Anatomical Structures.

Whole-organ plastic resin casting is a very useful method for preserving rare pathological specimens for forensic/anatomical studies and for teaching/research purposes. Many techniques have been proposed over time, but most of them use special non-commercially available resin mixtures, lengthy protocols, and are overall not easily implemented in any anatomy/pathology department that might need such a procedure for rapid organ preservation. Here, we utilized anatomical sections of the human brain, heart, kidneys, spleen, large intestine, and lungs from on-display organs that were fixed for more than 1 year in 10% neutral-buffered formalin and from a freshly processed cadaver for teaching purposes in our Human Anatomy Department, and we optimized a fast-processing protocol without the use of any clearing agents, which yields solid, clear, cylindrical resin casting blocks. The resulting protocol, which takes no longer than 4 days, proves that at least three commonly used epoxy resins from hobby shops can be utilized without any restrictions, and the use of resin or glycerin vacuum-forced impregnation even offers two choices of intrinsic contrast, depending on the nature of the preparation. A number of innovations have been included here and compared to existing publications, such as the use of a system of permanent fixation plexiglas rods that maintain the organ in the desired position and become invisible in the final block, the use of UVC sterilization of the tissue to ensure a long shelf life of the block, and the utilization of cheap cylindrical polypropylene food containers as casting molds. Altogether, we present a simple resin-embedding protocol that can be made available to any department/institution without the need for expensive materials and specially trained personnel.

Assessment of the Impact of Alternative Fixatives on HER2 Detection in Breast Cancer and Gastric Cancer Tumor Specimens.

The type of fixative used for preserving tumor specimens can significantly impact the performance of the immunohistochemistry and in situ hybridization assays used for assessing human epidermal growth factor receptor 2 (HER2) status. This study reports the prevalence of the use of alternative fixatives other than the guideline-recommended 10% neutral buffered formalin (NBF) during HER2 testing in a real-world setting. The effects of alternative fixatives [20% NBF and 10% unbuffered formalin (UBF) fixatives] on HER2 testing of breast cancer (BC) and gastric cancer (GC) cell lines and tissues are also assessed. Overall, 117,636 tumor samples received at a central laboratory from >8000 clinical trial sites across 60 countries were reviewed to determine the prevalence of alternative fixative usage. To investigate the impact of alternative fixatives, 27 cell lines (21 BC and 6 GC) and 76 tumor tissue samples (50 BC and 26 GC) were fixed in 10% NBF, 20% NBF, or 10% UBF, and evaluated for HER2 status by immunohistochemistry and in situ hybridization. Real-world data showed that 9195 (7.8%) tumor samples were preserved using an alternative fixative. In cell lines, overall percentage agreement, negative percentage agreement, and positive percentage agreement among the 3 fixatives were 100%. In tumor tissues, the agreement among 10% NBF, 20% NBF, and 10% UBF ranged between 94.7% and 96.6% for negative percentage agreement and 90.9% for overall percentage agreement compared with a range of 58.3% to 66.7% for positive percentage agreement. These results suggest that alternative fixatives may have the potential to convert HER2 status in tissues from positive to negative.

Equine herpesvirus myeloencephalopathy

Clinical History: A euthanized, 14-year-old, Warmblood gelding that had participated in an equine show was presented for necropsy and diagnostic workup to the San Bernardino laboratory of the California Animal Health and Food Safety Laboratory System. The animal had a 4-day history of mildly swollen limbs and hyperthermia, and developed neurologic signs shortly before euthanasia. Necropsy Findings: The urinary bladder had multifocal to coalescing hemorrhages in the mucosa and approximately 5 ml of turbid urine with sandy sludge (Figure 1). The entire spinal cord was removed and cross-sectioned serially after fixation in 10% neutral-buffered formalin during 48h. Multifocally, in multiple sections of the cervical, thoracic, and lumbar segments, there were uni- or bilateral and asymmetrical, wedge-shaped areas of gray discoloration and hemorrhage (Figures 2-4). In addition, there were extensive hemorrhages around the nerve roots of the cauda equina.

A practical protocol to prepare paraffin-embedded whole tick histology sections

Ticks are important ectoparasites that are capable of transmitting multiple classes of pathogens and are currently linked with many emerging tick-borne diseases worldwide. With increasing occurrences of tick-borne diseases in both humans and veterinary species, there is a continuous need to further our understanding of ticks and the pathogens they transmit. Whole tick histology provides a full scope of the tick internal anatomy, allowing researchers to examine multiple organs of interest in a single section. This is in contrast to other techniques that are more commonly utilized in tick-borne disease research, such as electron microscopy and light microscopy of individual organs. There is a lack of literature describing a practical technique to process whole tick histologic sections. Therefore, the current study aims to provide researchers with a workable protocol to prepare high quality paraffin-embedded whole tick histology sections. Amblyomma americanum adults were used as an example species for this study. After a series of pilot experiments using a combination of various fixatives, softening agents and processing techniques, we elected to compare two common fixatives, 10% neutral-buffered formalin (NBF) and Bouin's solution for whole ticks. Equal numbers of A. americanum unfed adults (n = 10/fixative) were processed identically and their whole tick histology coronal sections were individually scored. Higher scores were assigned to whole tick sections that contained more internal organs that are crucial for tick-borne disease research (e.g. salivary glands and midgut), high integrity of tissues and exoskeleton on the section, and good fixation and staining quality of the tissues. The mean total scores for Bouin's-fixed ticks were significantly higher compared to NBF-fixed ticks (p = 0.001). To further assess our preferred technique, we also demonstrated the feasibility of producing high quality whole tick sections for three other common tick species of medical importance (Rhipicephalus sanguineus, Ixodes scapularis, and Dermacentor variabilis) using Bouin's solution. While this technique may require further optimization for other tick species, we described a feasible protocol that uses commonly available tools, reagents and standard histologic equipment. This should allow any investigator to easily make adjustments to this protocol as needed based on their experimental goals.

Growth, pectoralis muscle performance, and testis of pelung cockerels (Gallus gallus gallus [Linnaeus, 1758]) supplemented with blood clam shell powder (Anadara granosa [Linnaeus, 1758]).

Pelung cockerels (Gallus gallus gallus [Linnaeus, 1758]) are different from other native cockerels in that they have a long and unique voice, in addition to their tall, large, and sturdy body with a relatively heavy body weight (BW). The sound quality of pelung cockerels is affected by the structure of the syrinx and their large and strong chest muscles. The performance of the chest muscles, and subsequently its voice, is influenced by the hormone testosterone. The shell of blood clams (Anadara granosa Linnaeus, 1758), a saltwater bivalve is known to contain a natural aromatase blocker (NAB) capable of blocking the aromatase enzyme from converting testosterone to estradiol. This generates consistently high levels of testosterone. This study aimed to determine the effect of blood clam shell powder (BCSP) as an NAB on the growth, pectoralis muscle performance, and testes of pelung cockerels. The study design was a completely randomized design, with 16 pelung cockerels aged 40-56 weeks divided into four treatment groups: T0 (control); T1 (BCSP [A. granosa] 0.9 mg/kg BW); T2 (zinc sulfate [ZnSO4] 0.9 mg/kg BW); and T3 (testosterone 3 mg/day). The animals were acclimatized for 7 days and then given dietary treatments for 56 days. The measurement of the comb, wattle, and chest circumference (CC) of pelung cockerels was performed on days 0, 14, 28, 42, and 56. At the end of the treatment, the pelung cockerels were sacrificed and the data of the pectoralis muscle weight (PMW), testis weight (TW), and area of the pectoralis muscle (APM) were measured. Samples of pectoralis muscle and testes were taken and fixed in 10% neutral buffer formalin for histology. The proliferating cell nuclear antigen (PCNA) was identified by immunohistochemical staining. To measure fascicle area (FA), myofiber area (MA), and enumerate, the fascicle myofibers (NM) histology preparations were stained with hematoxylin and eosin (H and E). Testicular preparations were stained with H and E to measure the diameter of the seminiferous tubules (DST) using ImageJ software. The growth performance on day 56 showed significantly (p < 0.05) higher differences of CC in T1 compared to T2 and T0, in T1 and T3 compared to T0, and in T3 and T2 compared to T0. Pectoralis muscle results, that is, FA, NM, MA, and PCNA-positive cells, showed that cockerels on treatment T3 had significantly higher results than other treatments, T1 was significantly different from T2 and T0, and T2 was significantly different from T0. In addition, the TW and DST measurement of cockerels on treatment T3 were significantly reduced (p < 0.05) than the other treatment groups. The oral administration of BCSP in the role of a NAB at a dose of 0.9 mg/kg BW for 56 days improved the growth performance and pectoralis muscle, especially the CC, FA, NM, MA, and PCNA-positive cells parameters, but did not affect the PMW, APM, and testis of pelung cockerels. The administration of testosterone at 3 mg/day for 56 days contributed to the decrease in TW and DST, as well as atrophy of the seminiferous tubules of pelung cockerels.

The Application of Guanidinium to Improve Biomolecule Quality in Fixed, Paraffin-embedded Tissue.

Neutral buffered formalin (NBF) is the most common fixative in clinical applications. However, NBF damages proteins and nucleic acids, limiting the quality of proteomic and nucleic acid-based assays. Prior studies have demonstrated that BE70, a fixative of buffered 70% ethanol, has many benefits over NBF but the degradation of proteins and nucleic acids in archival paraffin blocks remain a challenge. Thus, we evaluated the addition of guanidinium salts to BE70 with the hypothesis that this may protect RNA and protein. Guanidinium salt supplemented BE70 (BE70G)-fixed tissue is comparable with that of BE70 via histology and immunohistochemistry. Western blot analysis also revealed that HSP70, AKT, and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) expression signals in BE70G-fixed tissue were higher than those in BE70-fixed tissue. The quality of nucleic acids extracted from BE70G-fixed, paraffin-embedded tissue was also superior, and BE70G provides improved protein and RNA quality at shorter fixation times than its predecessors. The degradation of proteins, AKT and GAPDH, in archival tissue blocks is also decreased with the addition of guanidinium salt to BE70. In conclusion, BE70G fixative improves the quality of molecular analysis with more rapid fixation of tissue and enhanced long-term storage of paraffin blocks at room temperature for evaluation of protein epitopes.

Suitability of transbronchial needle aspiration for genotyping peripheral pulmonary tumors.

Transbronchial needle aspiration (TBNA) is a sampling tool that has demonstrated a higher accuracy in the diagnosis of peripheral pulmonary lesions (PPL) compared to other techniques. However, there are no studies investigating the value of TBNA in defining the genotype of peripheral lung cancer. To evaluate the accuracy of TBNA in defining the molecular characteristics of peripheral lung cancer. Consecutive patients who underwent TBNA for the diagnosis of a PPL at the Pulmonary Unit of the Azienda Ospedali Riuniti of Ancona (Italy) between January 2020 and September 2022 were included in the study. TBNA was performed under fluoroscopic guidance and the additional support of an ultrasound miniprobe, with an ultrathin bronchoscope with a flexible 21G needle. Samples were smeared on glass slides for cytological evaluation and flushed in 10% neutral-buffered formalin for cell-blocks. 154 patients were enrolled:55 were diagnosed with adenocarcinoma and 21 with squamous cell carcinoma. TBNA correctly diagnosed 43/55 (78.2%) patients with adenocarcinoma and 17/21 (81.0%) patients with squamous cell carcinoma, with a sensitivity of 77.5%. Complete genotyping for guiding targeted therapies was obtained in 52 patients (86.6%). TBNA is a valid tool for the diagnosis of PPL, allowing a correct diagnosis and a complete genotyping of the tumors in a considerable proportion of patients.

The Possible Effects of Vitamin D3 on AlCl3-Induced Histological and Morphometric Alterations of Adult Male Albino Rat Hippocampus.

Alzheimer's disease (AD) is a challenging neurodegenerative disease, and Vitamin D was proved to have neuroprotective effects. This study was conducted to evaluate the potential neuroprotective effects of Vitamin D3 supplementation on AlCl3-induced AD rat model in different hippocampal subregions (CA1, CA2, and CA3). It also aimed to compare the protective effects of protective versus therapeutic effects of Vitamin D3 regiments on the number of degenerated neurons and the neuronal layer thickness. Twenty-four adult male Albino Wister rats were sorted into GI: control; GII: AlCl3-AD model (100 mg/kg) orally for 42 days; GIII: Rats were co-treated with AlCl3 (as GII) and Vitamin D3 (400 IU/kg/day) orally for 42 days; GIV: Rats were treated with AlCl3 for 42 days then with Vitamin D3 for further 2 weeks. Sagittal sections (5 μ) from paraffin-processed brains previously fixed in 10% neutral-buffered formalin were stained with hematoxylin and eosin to evaluate the thickness and number of degenerated neurons in the hippocampal CA1, CA2, and CA3 subregions. The results of this study were expressed as mean ± standard deviation and analyzed by using IBM SPSS Statistics for Windows, version 23 (IBM SPSS, IBM Corp., Armonk, N.Y., USA). P < 0.05 was considered statistically significant. Vitamin D3 supplementations modulated the degenerative changes observed in the hippocampus of AD rat model. In all hippocampal subregions, the thickness was higher in rats treated with Vitamin D3 after the AD induction than rats treated with Vitamin D3 during AD induction. However, this increase was only significant in CA2. Comparison of the number of degenerated neurons between both groups treated with Vitamin D3 revealed that in CA1, the number of degenerated neurons did not statistically differ between the two groups. However, it was insignificantly lower in CA2 in rats treated with Vitamin D3 after the AD induction, and in CA3, it was insignificantly lower in rats treated with Vitamin D3 during the AD induction. Vitamin D3 was found to be effective in ameliorating histological and morphometric alterations in AlCl3-induced AD in rat model and could be proposed as both preventive and therapeutic supplements in high-risk AD patients.

Implementation of microsatellite instability testing for the assessment of solid tumors in clinical practice.

In Japan, microsatellite instability (MSI) testing for solid tumors was introduced in clinical practice in December 2018. Although immune checkpoint inhibitors (ICIs) are established standards of care for patients with MSI-high tumors, the status of implementing MSI testing in clinical practice remains unclear. We retrospectively reviewed the medical records of patients with solid tumors who underwent MSI testing between January 2019 and December 2020 at our institution. In total, 1,052 MSI tests were performed in 1,047 patients. Regardless of specimen volume and condition, the MSI status was successfully determined in 1,041 (99.0%) tests, encompassing 27 tumor types (microsatellite stable [MSS] or MSI-low: n=991 [95.2%] and MSI-high: n=50 [4.8%]). Patients whose specimens were fixed with 20% neutral buffered formalin (NBF) and who had specimens with prolonged storage (98.4% and 95.4%) showed lower success rates than those whose specimens were fixed with 10% NBF and who had specimens with nonprolonged storage (100.0% and 99.6%), respectively. The prolonged turnaround time (TAT) in MSI-high cases (median TAT: 24 days) was a critical issue that directly resulted in treatment delay. Of the 50 patients with MSI-high tumors, 24 (48.0%) received ICIs and 34 (68.0%) were referred to the Department of Clinical Genetic Oncology where 6 (12.0%) patients were diagnosed with Lynch syndrome. MSI testing was successfully performed for various types of tumors and specimens in clinical practice. Our study results identified certain issues associated with the clinical implementation of MSI testing, including optimal specimen selection, extended TAT in MSI-high cases, and awareness of hereditary tumors.

Liver Tissue Examination of Mice Using 10% BNF Fixation For 6 Hours And 16 Hours

The fixation process is the first stage in the manufacture of histopathological preparations which aims to preserve the tissue and harden the tissue, so that the tissue to be observed does not change in shape or size. BNF fixation solution (Neutral Buffer Formalin) 10% has been used as a routine fixative and It has been be the gold standard in histology laboratories for decades. The material used in this study was a 10% BNF solution because it is easier to use and it can be used to preserve tissue for a long period of time. This study aims to determine the differences in the microscopic results of the liver tissue of mice (Mus musculus) fixed with 10% BNF for 6 hours and 16 hours. This type of research is descriptive analytic. The research was carried out at the Barokah Laboratory/ dr.Mezfi Unita on 04-05 March 2022 with a total sample of 20 samples of mice liver tissue (Mus musculus). The results of the study on 10% BNF fixation for 6 hours and 16 hours showed that the average results were not good. The conclusion from the results of the study that there was no difference in the microscopic results of liver tissue that were fixed with 10% BNF for 6 hours and 16 hours using Hematorylin-Eosin staining. 10% BNF fixation with a time of 6 hours and 16 hours can be used as an alternative in histopathological examination in the short term and long term.

Cytoarchitecture of greater cane rat (Thryonomys swinderianus) habenulae and mammillary body

This study was conducted to investigate expression of neurons, glia and connective tissue markers in the habenulae and mammillary body of the Greater Cane Rat (GCR) using histological and immunohistochemical techniques. Four apparently healthy males GCR obtained from Lagos Nigeria were used for this experiment. The brains were harvested after perfusion, carefully dissected and post-fixed in 10% neutral-buffered-formalin (NBF) at 40C for 48 hours. Subsequently, tissue sections were prepared and stained using Haematoxylin and Eosin (H&amp;E), Cresyl-violet, ionized calcium-binding-adapter molecule 1 (Iba1; microglia), glial fibrillary acidic protein (GFAP; astrocytes), neuronal nuclei (NeuN; neurons) and collagen type 1 (Collagen1) antibodies. H&amp;E and Cresyl-violet staining showed the neuronal nuclei and their fibres in the habenulae, mammillary body, and pial capsule that surround the mammillary body ventrally. Astrocytes in the habenulae showed positive immunoreaction to anti-GFAP while the mammillary body revealed expression of perivascular astrocytes and capsular astrocytes. Also, in the mammillary body, the perivascular microglia and meningeal microglia in the pial capsule showed immunopositive reaction to anti-Iba1 while the fibres within blood vessels and capsule were immunopositive to collagen type 1 antibody in the same structure. This work has provided the first cytoarchitectural features and clear description of expression of these specific markers in habenulae and mammillary body in the GCR, which are similar to other rodents but with slight variations.

Frozen Versus Paraffin-Embedded Tissue for Glomerular Basement Membrane Thickness Measurement

In kidney pathology, quantification of glomerular basement membrane (GBM) thickness is a useful marker of several kidney diseases. The standard technique to measure GBM thickness is transmission electron microscopy (TEM). In our laboratory, renal cortex is routinely submitted for TEM evaluation in a neutral-buffered formalin solution. Frozen tissue or paraffin-embedded tissue is reclaimed when wet tissue is not available for TEM or when glomeruli cannot be found in the submitted formalin-fixed tissue.