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AIM To assess the morphological and functional characteristics of spermatozoa and DNA stability in patients with normal seed parameters and with an ITZ exceeding the threshold value. PATIENTS AND METHODS 85 males were studied for semen quality parameters. The morphological anomalies of head, middle piece, and tail of spermatozoa were estimated and presented in illustrations. Due to results elaborated the index of teratozoospermia was calculated. Then the Comet assay was performed. RESULTS It has been found that spermatozoa from patients with high teratozoospermia index possessed decreased motility as compared to ones from patients with low index of teratozoospermia. It was not found any difference in concentration of spermatozoa between groups of males which possess low and high index of teratozoospermia. The spontaneous frequency of DNA damage estimated by means of single cell gel electrophoresis followed by silver staining in spermatozoa from healthy donors did not exceed 3,1+/-0,6. The frequency of DNA damages in sperm of patients with abnormal index of teratozoospermia exceeded 39,6+/-7,4% (p

A 6‐year‐old female bottlenose dolphin (Tursiops truncatus) kept in dolphinarium died after a 3.5‐month period of lethargy and inappetence despite antibiotics and supportive care. At necropsy, gross findings included diffuse varying‐sized nodules in the lungs and scattered nodules throughout the heart, spleen, mesenteric and hilar lymph node and kidney. Microscopically, the lesions were characterised by disseminated fungal pyogranulomas with numerous intralesional Mucor‐like fungi. The fungi structures were demonstrated by Periodic acid‐Schiff and Gomori methenamine silver stain. Molecular analyses of the fungi were Rhizopus microsporus by PCR sequencing 18S ribosomal RNA gene. Ziehl–Neelsen stain failed to show acid‐fast bacterial infection. Based on pathological and molecular examination, systemic granulomatous mucormycosis was diagnosed. To our knowledge, this is the first reported case of systemic mucormycosis caused by Rhizopus microsporus in bottlenose dolphin., This study reported systemic mucormycosis in a bottlenose dolphin (Tursiops truncatus). The presence of the Rhizopus microsporus in bottlenose dolphin was reported for the first time in China.

Electrophoresis is one of the major techniques to analyze macromolecular structure and interaction. Its capability depends on the sensitivity and specificity of the staining methods. We have here examined silver staining of proteins and nucleic acids separated by agarose native gel electrophoresis. By comparing five commercial kits, we identified Silver Stain Plus from Bio-Rad most adequate, as it provided little background staining and reasonable band staining. One of the disadvantages of the Silver Stain Plus kit is its variable staining of glycoproteins as tested with several model samples, including hen egg white proteins, α1-acid glycoprotein and SARS-CoV-2 Spike protein. One of the advantages of silver staining is its ability to stain nucleic acids as demonstrated here for a model nucleic acid with two kits. It was then used to monitor the removal of nucleic acids from the affinity-purified maltose binding protein and monoclonal antibody. It also worked well on staining proteins on agarose gels prepared in the vertical mode, although preparation of the vertical agarose gels required technological modifications described in this report. With the silver staining method optimized here, it should be possible in the future to analyze biological samples that may be available in limited quantity.

The current study investigated the morphological, histochemical, and immunohistochemical characteristics of alarm cells and their precursors in ruby-red-fin shark (rainbow shark), Epalzeorhynchos frenatum (Teleostei: Cyprinidae). Precursor alarm cells were shown to be small, cuboidal, pyramidal, or round in shape, with eosinophilic cytoplasm, resting on the basement membrane of the epidermis. The cells later elongated to become columnar in shape. Subsequently, they enlarged and became large oval-shaped cells. They then underwent shrinkage and vacuolation. The superficial alarm cells were collapsed. Alarm cells were found to have an affinity for different histochemical stains, including bromophenol blue, iron hematoxylin, Sudan black, Mallory triple trichrome, Crossman's trichrome, Safranin O, and Weigert's stains, as well as lipase and alkaline phosphatase. Endocrine properties of the alarm cells were identified by silver staining and synaptophysin immunostaining. Alarm cells exhibited stemness activities and had a strong immunoaffinity for CD117. They had a proteolytic function, as identified by lysosome-specific staining with acridine orange and strong immunoaffinity for matrix metalloproteinase (MMP-9). They also exhibited proliferatively, reflected by immunological staining by proliferating cell nuclear antigen. In conclusion, alarm cells are unique epidermal cells with multiple functions. They play immunological, and endocrine, roles. They also retain stemness and proliferative properties.

The interaction of explosion-induced blast waves with the head (i.e., a direct mechanism) or with the torso (i.e., an indirect mechanism) presumably causes traumatic brain injury. However, the understanding of the potential role of each mechanism in causing this injury is still limited. To address this knowledge gap, we characterized the changes in the brain tissue of rats resulting from the direct and indirect mechanisms at 24 h following blast exposure. To this end, we conducted separate blast-wave exposures on rats in a shock tube at an incident overpressure of 130 kPa, while using whole-body, head-only, and torso-only configurations to delineate each mechanism. Then, we performed histopathological (silver staining) and immunohistochemical (GFAP, Iba-1, and NeuN staining) analyses to evaluate brain-tissue changes resulting from each mechanism. Compared to controls, our results showed no significant changes in torso-only-exposed rats. In contrast, we observed significant changes in whole-body-exposed (GFAP and silver staining) and head-only-exposed rats (silver staining). In addition, our analyses showed that a head-only exposure causes changes similar to those observed for a whole-body exposure, provided the exposure conditions are similar. In conclusion, our results suggest that the direct mechanism is the major contributor to blast-induced changes in brain tissues.

Aim. To study the activity of nucleolar organizer regions (NORs) in different Ukrainian cattle breeds in terms of their apparent activity status in silver stain and possible relation with milk productivity. Methods. Chromosome prepara- tions using lymphocytes from the peripheral blood of 90 cows of different breeds were used in the study. NOR activity was determined by visual evaluation of concentrations of silver precipitation on NORs in individual chromosomes. A 50 % silver nitrate solution was used to stain chromosome preparations. NORs were detected as dark spots on telomeres of the corresponding chromosomes. Results. The cytological analysis of chromosome preparations from lymphocytes of first lactation cows detected NOR polymorphism in Ukrainian Red-and-Motley dairy cattle (URM), Ukrainian Black-and-White dairy breed (UBW), and hybrid cows, obtained by crossing Ukrainian Red-and-Motley dairy breed and Montbeliarde bulls (URM × M). First lactation cows of URM and UBM had higher or the same inci- dence of cells with four (29.8 and 30 %) and five (17.1 and 19.5 %) NORs, while in URM × M cows the incidence of cells with the same number of NORs was almost twice lower; cells with 7 and 8 NORs exceeded a similar index for other investigated breeds almost twice (2.5 against 4.5 % and 2.0 against 4.2 %). The highest level of chromosomal aberrations (CA) was observed in the group of animals with medium number (2 to 3 NORs per cell), and the lowest – in the group with a high number of NORs (from 6 to 7) with a reliable intergroup difference (p < 0.01). NOR activity was the highest in the group of animals of local origin (URM × M) with a milk yield over 7,000 kg in 305 days of the first lactation and the lowest in the UBW cows with a milk yield of 4–5,000 kg during the first lactation. Conclusions. We determined the differences in the activity of nucleolar organizers between the investigated groups of cows of dairy breed. URM × M hybrids reliably (р ≤ 0.05) exceeded dairy UBW cows by this index. No statistically significant different was found between other investigated groups of animals by this trait. Higher dairy productivity was found in the animals with higher frequency of NORs in the chromosomes of metaphase cells. In our opinion, the number of active NORs demonstrates relative variability between their number and the rate of protein synthesis, required to implement the productivity traits of the investigated animals.

In July 2020, a sow in a breeding herd in the Chiba Prefecture, Japan, suffered abortion. A necropsy revealed pale pulmonary foci scattered in the two fetuses. Histologically, multifocal pulmonary necrosis was detected with numerous yeasts. The yeast was positively stained using the periodic acid-Schiff reaction and Grocott's silver stain. Molecular identification indicated that the yeast was Candida parapsilosis. In conclusion, our results suggested that C. parapsilosis caused multifocal necrotizing pneumonia in the two fetuses. This study is the first report of a swine abortion with C. parapsilosis infection.

Background Pneumocystis pneumonia (PCP) is a fatal infectious disease caused by Pneumocystis jirovecii (PJP). The major factor relevant to morbidity and mortality seems to be the host inflammatory reaction. The objective of this study was to evaluate the role of IL-2, IL-4, IL-10, and IL-13 cytokine mRNA expression among suspected P. jirovecii infection. Methods This was a cross-sectional analytical study undertaken in Aseer region, Saudi Arabia. One hundred suspected PCP cases and 100 healthy controls were included in the study. Basic clinical manifestations, radiological findings, microbiological and immunological findings were extracted from the hospital records from January 2019 to August 2019, Pneumocystis detection was done by immune-fluorescent staining (IFAT, Gomorimethanamine silver staining (GMSS), Giemsa staining, Toluidine blue O (TBO), and Pneumocystis RT-PCR. Results Increased more than 5 fold, 3 fold, 4 fold, and 7 fold of IL-2, IL-4, IL-10, and IL-13 mRNA expression were observed in PCP cases compared to controls. Higher expression of IL-2 mRNA was connected with crept, wheezing and chest X-ray findings like central perihilar infiltrate, patchy infiltrate, consolidation, hilar lymphadenopathy, pneumothorax, pleural effusion which showed higher expression compared to counterpart (pp=0.002), dyspnea (p=0.003), crept (p=0.01), and chest X-ray findings (p Conclusion Confirmed cases of P. jirovecii showed higher IL-2, IL-4, IL-10, and IL-13 mRNA expression comparatively to negative cases. Increased expression of cytokines may be indicative of infection severity and could help in patients’ management.

The samples used for forensic DNA analysis in living individuals are usually blood and buccal swabs, however, blood collection requires an invasive method that can cause discomfort, thus a buccal swab can be a good choice for individuals examined, especially children. This study aimed to determine the effect of temperature and storage time of buccal swabs on the quantity of DNA as material for DNA examination in the forensic field. This study was a laboratory experiment to determine the effect after treatment. Buccal swab samples were 48 and divided into 2 temperature groups, namely room temperature (RT) and 4℃. The division of the temperature groups was also observed with time differences, namely 1, 3, 5, 7 days. EDNA extraction used the DNAzol method and DNA quantification used a Spectrophotometer. The PCR process was carried out with STR primers FGA and D13S317 loci. The visualization stage used acrylamide gel and silver staining. The results of this study prove that there is an effect of temperature and storage time of buccal swab samples. The longer the treatment time, the lower the DNA level. With statistical analysis, it is obtained p-value of

The scuticociliatid ciliates Ancistrum haliotis and A. crassum are parasites that may cause high mortality in the cultured abalone Haliotis spp. and the bivalve Ruditapes philippinarum. Traditional identification with silver staining methods is hampered by their morphological similarities to closely related species and the complicated procedures of morphological analysis. We designed two SSU rRNA-targeted oligonucleotide probes labeled with a fluorochrome, and optimized the fluorescence in situ hybridization (FISH) protocols for identification of A. halioti and A. crassum, respectively. The assays resulted in a clear identification by strong fluorescence signals from the oligonucleotide probes. The method can be used for quick and accurate quantitative analysis of A. haliotis and A. crassum infections on host molluscs.

(1) What are the structures indicated by the blue arrows?(2) In this silver stain, which step/reagent is omitted that would normally prevent the yellow background staining?(3) What disease often pr...

Introduction: This experimental study gauged the value of argyrophilic nucleolar organizer region (AgNOR) staining as a possible technique for the estimation of cell kinetics in conventional histology sections, in benign and malignant breast lesions. Methods: With a silver staining technique and immunohistochemistry, we associated the numbers of AgNORs and Ki67 scores in 30 breast carcinomas and 10 benign breast lesions. Results: The mean values of Ag NORs silver stain dots count for normal, benign, grade II and III were 1.28±0.17, 2.83±0.68, 5.23±0.87and 7.32±0.92 respectively. There was a statistically significant difference (p≤0.001) was noticed between the all individual groups, among the normal and breast lesion as well as among the GII and GIII. Immunohistochemical Results of Ki-67 protein exhibited homogenous golden-brown color in control case and a positive brown granules or diffuse dark brown color in the nuclei of both benign and malignant cases under the 400X magnify examined under the light microscope. Discussion: AgNOR counts performed on routine formalin-fixed paraffin sections could provide substantial kinetic evidence. Additionally, the difference in AgNOR counts between benign and malignant tumors is such that they may be of diagnostic worth., {"references":["Cowdry EV. Cancer cells. Philadelphia: Saunders Co; 1955.","Thompson CB. Apoptosis in the pathogenesis and treatment of disease. Science. 1995;267(5203):1456-62. doi: 10.1126/science.7878464.","Kerr JF, Winterford CM, Harmon BV. Apoptosis. Its significance in cancer and cancer therapy. Cancer. 1994;73(8):2013-26. doi: 10.1002/1097-0142(19940415)73:83.0.co;2-j.","World Health Organization (WHO). Breast cancer: prevention and control. 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Human papillomavirus (HPV) is believed to cause cervical cancer. Thousands of women develop cancer and other diseases caused by HPV each year. HPV 16 and 18 types are found in approximately 70% of cervical cancers. Micronuclei are small chromosomal fragments that are considered indicators of DNA damage. AgNOR positive dots are useful for assessing proliferation. We investigated the relation between HPV-DNA, micronuclei and AgNOR in smear samples. Three groups were defined: HPV negative, 16/18 positive and other high-risk groups (31, 33, 35, 39, 45, 51, 52, 56, 58, 66 and 68) (HR). After typing, micronuclei were identified by Papanicolaou staining and AgNOR regions were detected by silver staining. Serum reactive protein (CRP) also was measured. We found that the average age of HPV negative patients was significantly greater than for the HPV positive groups. We also found that CRP levels were significantly higher in the HPV 16/18 positive group than HPV negative and other HPV group. We found that the number of micronuclei in the HPV 16/18 group was significantly greater than for the HPV negative group. Also, we found that AgNOR staining for the HPV 16/18 group was significantly greater than for the HPV negative group. We found that CRP level, cell proliferation and genome instability were increased in HPV positive patients. The AgNOR and micronucleus tests were useful for evaluating cell proliferation and DNA damage.

Traumatic brain injury (TBI) causes substantial mortality and long-term disability worldwide. TGFβ1 is a unique molecular and functional signature in microglia, but the role of TGFβ1 in TBI is not clear. The purpose of this study was to investigate the role of TGFβ1 in TBI. The weight dropping device was used to establish TBI model of rats. Hematoxylin eosin staining and Bielschowsky silver staining were used to assess tissue loss. Beam walking and muscle strength tests were used to assess neurological deficits. Immunohistochemical staining was used to assess axonal injures. Western blotting was used to detect expression of related proteins. RT-PCR was used to detect expression of cytokines. Immunofluorescence staining was used to assess the microglia/macrophages activation. We observed obvious axonal injury and microglia/macrophages activation in the peri-lesion cortex. The expression of inflammatory cytokines was markedly high after TBI. The expression of TGFβ1 and TGFβRI were significantly reduced after TBI. TGFβ1 promoted the functional recovery and alleviated axonal injury 1 day after TBI. TGFβ1 promoted microglia/macrophages polarizing to alternative activation and alleviated neuroinflammation. These effects of TGFβ1 could be inhibited by LY2109761, the inhibitor of TGFRI/II. These results suggested that TGFβ1 played a protective role in axonal injury and could be a potential therapeutic target in early stages following TBI.

The current study investigates the polymorphic patterns of keratin-associated protein (KAP) 3.2 gene in Sandyno breed of sheep. Genomic DNA was isolated from blood samples of 51 numbers of Sandyno breed. Ovine specific primer associated PCR amplification of KAP 3.2 gene revealed product at 393 bp and genotyped by PCR-SSCP (Single Strand Conformation Polymorphism) method and visualized under silver staining technique. KAP 3.2 gene locus revealed 3 genotypes, viz. AA, AB and BB with a frequency of 0.84, 0.16 and 0 in Sandyno breed with allele frequencies of A(0.92) and B(0.08). Regarding population genetic indices, the effective number of alleles (Ne) for KAP 3.2 in Sandyno breed of sheep was found to be 1.1716. The PIC values was 0.1356 and FIS values was negative (– 0.0864) in this breed. The result revealed that the selected population of Sandyno breed of sheep was in Hardy-Weinberg equilibrium without any significant deviation from the population mean and was monomorphic for KAP 3.2 gene.

We report a 74-year-old male presented to an outpatient dermatology clinic in Manaus, Amazonas, with a one-year history of pruritic, keloidal lesions on his left lower extremity. Histopathology showed round structures in reticular dermis. Grocott methenamine silver stain revealed numerous round yeasts with thick double walls, occurring singly or in chains connected by tubular projections. The diagnosis was lobomycosis. Although the keloidal lesions presented by this patient are typical of lobomycosis, their linear distribution along the left lower limb is unusual.

Protein detection and quantification is a crucial part of many clinical, research, and preparatory applications [1]. Protein biomarkers, such as prostate-specific antigen (PSA) [2] and carcinoembryonic antigen (CEA), are regularly detected and quantified for cancer diagnosis and treatment monitoring [3]. On the other hand, cardiac troponin 1 (CTn1) is the most important protein to quantify to detect cardiac injury and myocardial infarction [4]. The detection of protein biomarkers such as PSA, CEA, and CTn1 requires specific identification and quantification of the protein, eliminating detection techniques that enable only size-based quantification of proteins in general, such as silver staining and simple gel electrophoresis [5].

A novel electrochemical gene detection method based on bio-barcode/gold label silver stain (GLSS) dual amplification is presented. The silica nanoparticle/gold nanoparticles (SiO2/AuNPs) barcode probes were constructed by combining SiO2 nanoparticle-labeled DNA with AuNPs, which hybridizes with target DNAs and the electrode surface-modified DNA probes. The silver ions were reduced on the surface of AuNPs to form a silver shell by the catalysis of AuNPs, and then the electrochemical detection of deposited silver was performed by linear sweep voltammetry (LSV). In this paper, the effects of electrode blocking agent, blocking time and silver deposition time on electrochemical detection were also investigated. The proposed electrochemical DNA biosensor bring forth a low detection limit of 0.23 fM and a widely linear range of 1 fM to 10 pM. This method can clearly distinguish the complementary and mismatched DNA, with good repeatability and stability.

Purpose Cat scratch disease (CSD) is an infectious disorder caused primarily by the bacterium Bartonella henselae (B. henselae). Immunohistochemistry (IHC) and Warthin-Starry silver stain (WS) are considered to be indispensable to diagnose CSD in combination with morphologic characteristics. In this study, we retrieved and reviewed 46 cases of paraffin-embedded lymphadenitis with histologic and/or clinical suspicion of CSD between 2014 and 2018, and detected B. henselae by IHC and WS, respectively, and evaluated the application significance of IHC and WS for the detection of B. henselae and validated their values in the pathologic diagnosis of CSD. Materials and methods B. henselae was detected by IHC and WS; validation of 2 methods for detecting B. henselae was evaluated by sensitivity, specificity, false-positive rate, false-negative rate, precision, negative predictive value, and agreement rate. Results Microscopically, suppurative granulomas and/or multiple stellate microabscesses were observed in the accessory cortex of lymph nodes, especially near the subcapsule. Our results showed that 80.4% (37/46) of cases were positive for B. henselae by IHC, manifesting mainly punctuate, granular, or linear to outline the shape of bacteria. However, the positive rate of B. henselae by the WS method was 52.2% (24/46). There was a significant difference between IHC and WS (P=0.023). Moreover, a positive percentage of B. henselae was 97.8% (45/46), which was detected by the combined application of IHC and WS. The combination of IHC and WS exhibited high sensitivity (97.8%) and good agreement rate (86.5%). Conclusion The combined application of the IHC and WS method may have important clinical advantages, which is with the highest sensitivity and agreement rate for pathologic diagnosis of CSD.

Context.—Direct visualization of Pneumocystis jiroveci organisms, using Gomori methenamine silver (GMS) staining in bronchoalveolar lavage fluid (BAL), is a historical gold standard that has been widely used for the diagnosis of P jiroveci pneumonia (PJP). However, the stain may be less sensitive in human immunodeficiency virus (HIV)–negative immunocompromised patients owing to a lower burden of organisms.Objectives.—To assess the sensitivity of the GMS stain on BAL fluid for the diagnosis of PJP in HIV-negative immunocompromised patients as compared to HIV-positive patients.Design.—We conducted a retrospective review from 2012 to 2018 to identify immunocompromised patients (≥18 years old) who underwent bronchoscopy with BAL GMS staining for the diagnosis of PJP. To assess for sensitivity, we sought to identify BAL GMS-positive cases and BAL GMS-negative cases of PJP. The BAL GMS-negative cases were categorized into proven and probable PJP.Results.—We identified 45 adult immunocompromised patients with proven and probable PJP, including 24 HIV-negative (11 BAL GMS-positive and 13 BAL GMS-negative) and 21 HIV-positive cases (all were BAL GMS-positive). The sensitivity of BAL GMS for the diagnosis of PJP in HIV-negative immunocompromised patients was 11 of 24 (46%) versus 21 of 21 (100%) in HIV-positive patients (CD4: median, 10 cells/mL; range, 3–300 cells/mL). Delayed or missed diagnoses were seen in 3 cases of BAL GMS-negative PJP. Re-examination of BAL GMS slides showed rare P jiroveci cysts in 1 case.Conclusions.—BAL GMS has poor sensitivity for PJP in HIV-negative immunocompromised patients. Using BAL GMS as a sole method for PJP may result in missed or delayed diagnoses in this population.

MicroRNAs (miRNAs) have played a vital role in the regulation of gene expression and have been considered as potential biomarker candidates for early cancer diagnosis. Rapid and sensitive detection of microRNAs is highly desired. Here, we present a new method to rapidly and sensitively determine microRNAs based on the technology of gold nanoparticle catalyzed silver staining enhancement. The new method involves the sandwich hybridization of a capture probe immobilized on a magnetic bead, a reporter probe assembled on gold nanoparticles and a miRNA target, catalytic silver precipitation by gold nanoparticles, magnetic collection of the enhanced sandwich complex, dissolution of the silver precipitation and stripping detection. Using the proposed method the microRNA-7a assay was successfully carried out in less than 70 min and the detection limit was as low as 15 fM. The proposed biosensor may hold great promise in biological monitoring of microRNAs.

Nucleic acid polyacrylamide gel electrophoresis (PAGE) has played a critical role in the identification and characterization of viroid RNAs. In addition, double PAGE has been a very efficient tool for the detection of viroids as it is sequence independent and is based on the viroid structure of covalently closed/circular molecules. sPAGE has been widely used for the identification of new viroids as well as a routine detection tool.

Emerging evidence indicates that bone mass is regulated by systemic energy balance. Temperature variations have profound effects on energy metabolism in animals. Multiple studies have shown that mild cold exposure can reduce bone mass by affecting bone remodeling. However, the in-depth mechanism behind this phenomenon remains unclear. Therefore, to address this issue, we exposed 2-month-old C57BL/6J male mice to cold (4 °C) and normal (23 °C) temperatures for 28 days and studied the effects of cold exposure on bone mass. Micro-computed tomography studies showed that the bone volume fraction was significantly reduced after 14 days of exposure to cold temperature. However, the same was recovered after 28 days. Ploton silver staining and immunohistochemical studies further revealed that exposure to cold decreased the canalicular length, number of E11- and MMP13-positive osteocytes after 14 days, but they returned to the baseline levels after 28 days unlike in the normal temperature control group. In addition, changes in Caspase-3 levels and osteocyte population indicated that exposure to cold temperature augmented apoptosis of osteocytes, while the same declined post 14 days and 28 days, respectively, conceivably affecting the bone mass directly. In vitro results confirmed the positive effect of brown adipocytes on osteocytes’ dendrites and E11 expression level. In conclusion, our findings indicate that exposure to cold temperature can influence bone mass in a time-dependent manner, with bone mass decreasing and recovering at 2 and 4 weeks, respectively. This fluctuation in bone mass may be evident due to the apoptosis and consequent decline of osteocytes.

Certain contemporary histology stains and methods are not the same as those used in the past. This progression has delved into the requirement for more precise, less complex, and efficient staining procedures. The objective of this study is to assess historical and contemporary stains and procedures, as well as the challenges surrounding their improvement. Carmine, hematoxylin, silver nitrate, Giemsa, trichome stain, Gram stain, and mauveine were among the first histological stains discovered in nature. Aside from their utility in the study of tissues at the time, they also laid the groundwork for the development of commercial dyes that are still in use today. Hematoxylin and eosin, Ziehl-Nielsen (ZN) stain, periodic acid-Schiff stain, and Grocott-Gomori methenamine silver stain are some of the most recently developed histological stains. The future of histological stains and processes appears to be influenced by technological advancements and the demand for cost-effective diagnostic approaches in the healthcare system. Thus, currently used histological stains appear to be economical, quick, and reliable tools for interpreting, archiving, and delivering essential diagnoses that could not be achieved by any other means.

Introduction Honey is known for exhibiting antibacterial properties, indicating its use as part of traditional medicine since the early ages. With the advent of antibiotic-resistant bacteria, the need for alternative antimicrobials has outpaced the actual development of novel, broad-spectrum antibiotics. Previous research has revolved around the sugar content of honey because its sweetness makes it an attractive food source. However, research assessing the protein and lipid components of honey is lagging behind that of its sugar counterpart. The goal of this investigation was to examine the antimicrobial properties of honey and to identify any distinct proteins or lipids. Methods In order to isolate individual peptides and lipids, the different samples of local and foreign-sourced honeys were dialyzed, and the resulting dialysate proteins were screened via gel electrophoresis (sodium dodecyl-sulfate polyacrylamide gel electrophoresis [SDS-PAGE]) with Coomassie blue and silver stain, while lipids were examined using thin layer chromatography (TLC). To assess antimicrobial potency, a series of Kirby-Bauer disc diffusion assays was performed on Mueller-Hinton agar using different types of raw honey with Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa, Escherichia coli, and Bacillus subtilis. The process was then repeated using the peptide extracts from the dialyzed fractions of the honeys. Results The SDS-PAGE trials revealed repetitive promising protein bands across several gels below 75kDa with both Coomassie blue and silver staining. The TLC analysis of varying raw honey samples consistently demonstrated the presence of medium and long-chain fatty acids, likely in the range of C12-C14. In the disc diffusion assays, the greatest amount of inhibition was seen when the honeys were tested as a whole instead of its constituent parts. Conclusion Instead of an individual component acting as the key to honey's action against bacteria, it appears there is a synergistic relationship amongst the sugars, proteins, and lipids that make each honey unique.

Pathology is practiced by visual inspection of histochemically stained tissue slides. While the hematoxylin and eosin (H&E) stain is most commonly used, special stains can provide additional contrast to different tissue components. Here, we demonstrate the utility of supervised learning-based computational stain transformation from H&E to special stains (Masson’s Trichrome, periodic acid-Schiff and Jones silver stain) using kidney needle core biopsy tissue sections. Based on the evaluation by three renal pathologists, followed by adjudication by a fourth pathologist, we show that the generation of virtual special stains from existing H&E images improves the diagnosis of several non-neoplastic kidney diseases, sampled from 58 unique subjects (P = 0.0095). A second study found that the quality of the computationally generated special stains was statistically equivalent to those which were histochemically stained. This stain-to-stain transformation framework can improve preliminary diagnoses when additional special stains are needed, also providing significant savings in time and cost., Performing multiple histological stains on a biopsy can be costly and time consuming. Here the authors present a method for the digital transformation of H&E stained tissue into special stains (e.g., PAS, Masson’s Trichrome and Jones silver stain), and demonstrate that it improves diagnoses over the use of H&E only.

Herein, we report a novel fluorogenic visualization method for mouse brain tissues by making use of a Ag+-specific aggregation-induced emission (AIE) strategy after silver impregnation. This novel fluorogenic silver-AIE method surpasses the chromogenic detection used by many conventional silver staining protocols, which have historically made revolutionary contributions to neuroanatomy. This fluorescent silver-based stain allows for analysis of images acquired using wide-field, confocal, SIMs, or light-sheet fluorescence microscopy. We validated the sensitivity, specificity, and versatility of the silver-AIE stain as compared to conventional silver stains and other neuron-specific ones. The silver-AIE stain is demonstrated to be highly compatible with immunofluorescence co-staining in paraffin sections and can be applied to hydrogel-based cleared tissue by the passive CLARITY method for image analysis of large tissue slices of 300 μm thick in this study. From this fluorogenic strategy, it is possible for vast types of classic silver staining methods to be modified and re-vitalized.

Synucleinopathies [Parkinson’s disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA)] share filamentous α-synuclein assemblies in nerve cells and glial cells. We compared the abilities of brain extracts from MSA and PD patients to induce neuronal α-synuclein assembly and neurodegeneration following intracerebral injection in heterozygous mice transgenic for human mutant A53T α-synuclein. MSA extracts were more potent than PD extracts in inducing α-synuclein assembly and in causing neurodegeneration. MSA assemblies were Campbell-Switzer- and Gallyas-silver-positive, whereas PD assemblies were only Campbell-Switzer-positive, in confirmation of previous findings. However, induced α-synuclein inclusions were invariably Campbell-Switzer-positive and Gallyas-negative, irrespective of whether MSA or PD brain extracts were injected. The α-synuclein inclusions of non-injected homozygous mice transgenic for A53T α-synuclein were also Campbell-Switzer-positive and Gallyas-negative. These findings demonstrate that transgene expression and its intracellular environment dominated over the silver staining properties of the conformers of assembled α-synuclein.

Introduction: Pap smear has been utilized since a long time in detecting cervical pre-cancers and cancers. Their potential in diagnosing pathogenic organisms however is underestimated. Reporting organisms morphologically consistent with Candida species, in cervicovaginal smears, is easier, but coexisting coccobacillary flora is often missed in the bargain. Such co-existing infections make the treatment difficult for clinicians, prolonging the distressing symptoms of patients. We evaluated modified silver staining technique using a microwave - JONES MARRES STAIN (JM) and tried comparing the results with conventional Pap smears to find any additional flora. Aim: To identify co-existing organisms in addition to Candida in a Pap smear stained with Jones Marres stain. Materials and methods: This retrospective prevalence study that included 82 consecutive conventional Pap smears routinely stained with Pap stain was then subjected to silver stain in a microwave. Inclusion criteria: Pap smears reported as showing Candida group of organisms. Results: In the 82 smears evaluated, JM stain helped in easier picking up of additional coccobacillary flora in 37(45%) of cases in comparison to 4(4.87%).These coexisting infections were missed in conventional Pap smear reporting. Conclusion: Modified JM stain helped in diagnosing additional flora thus reducing the chances of missing out co-infections and hence is useful for providing effective treatment Keywords: Microwave Silver stain, Co-infection, Pap smear, Coccobacillary flora.

A novel gold-label silver-stain electrochemical immunosensor was developed based on polythionine-gold nanoparticles (PTh-Au) modified glassy carbon electrode (GCE) as a platform and secondary antibody labeled Au NPs (Ab2-Au) as immumoprobe for carcinoembryonic antigen (CEA) detection. The sandwich-type biosensor adopted anodic stripping voltammetry to detect silver stripping signal when the Ab2-Au of the formed immunocomplexes were stained with silver. The optimized detection conditions were investigated. The effect of different electrochemical responses at various concentrations of CEA was checked by anodic stripping voltammetry. This immunosensor showed a low detection limit of 0.055 ng/mL and a wide linear calibration of 0.1–120 ng/mL (R2 = 0.99856). Moreover, this immunoassay also existed the advantages of good reproducibility, stability and selectivity. Thus, this immunosensing protocol may provide a potential application for effective clinical detection of CEA.

Nucleolar organiser regions (NORs) are loops of DNA which are responsible for ribosomal RNA (rRNA) transcription in the nucleus of cells. NORs can be selectively stained by silver methods. The NORs agyrophilia is due to a group of nucleolar proteins, which have affinity for silver (Agyrophilic Nucleolar Organizer Regions, AgNOR proteins). The aim of the present study is to assess proliferative index in different grades of invasive ductal carcinoma (IDC) by using AgNOR as a marker. A total of 18 formalin fixed paraffin-embedded breast tissue blocks were retrieved from the archives of the histopathology department of the Komfo Anokye Teaching Hospital, Ghana. The tissue blocks included 17 IDC, (with 8 as grade 2 IDC, 9 as grade 3 IDC) and 1 normal breast tissues. Tissue sections were diagnosed and graded following haematoxylin and eosin (H&E) staining. Silver staining was used to demonstrate the presence of AgNOR. The AgNOR dots were manually counted in 50 and 100 cells. The mean AgNOR count (mAgNOR) and proliferative index, (pAgNOR, percentage of cells with ≥5 AgNOR dots per nucleus), significantly increased with increasing grade of IDC. The mAgNOR and pAgNOR were low in normal breast, (at p-value less than 0.05). There was no significant difference in the mAgNOR and pAgNOR score for counting 50 and 100 cells (at a p-value of 0.01). mAgNOR and pAgOR increases with increase in grade of IDC and hence can be used to assess proliferative index of IDC. Keywords: Nucleolar organiser regions (NORs), Agyrophilic Nuclear Organizer Region (AgNOR), Proliferative activity, Breast Cancer

Polyacrylamide gel electrophoresis, followed by an appropriate staining, is a popular and useful analytical procedure for protein identification and characterization. The aim of this study was to develop a method for protein visualization in polyacrylamide gels that would be alternative to Coomassie blue or silver staining. The proposed method is simple, fast and inexpensive. The optimized protocol for protein staining and visualization takes as little as 6 minutes and utilizes deionized water and chloroform. Fluorescence of proteins is induced by UV light and can be detected with a standard transilluminator.