Biotechnic & Histochemistry最新文献

{"title":"Micronucleus cytome assay reveals genomic instability in premalignant cervical lesions among high-risk HPV-positive women.","authors":"Nohelia Cajas-Salazar, Leidy J Mueses-Moreno, Carlos H Sierra-Torres","doi":"10.1080/10520295.2026.2732168","DOIUrl":"https://doi.org/10.1080/10520295.2026.2732168","url":null,"abstract":"<p><p>Cervical cancer is a major health burden driven by persistent infection with high-risk human papillomavirus (HPV). Genomic instability biomarkers from the micronucleus cytome (MNcyt) assay may improve identification of women at risk of progression beyond cytology and HPV testing. A cross-sectional study of 184 women attending routine cervical cancer screening was conducted in Popayán, Colombia. Exfoliated cervical epithelial cells were collected for HPV DNA genotyping, liquid-based cytology, and MNcyt analysis using the Feulgen-Fast Green DNA-specific method. For each sample, 2,000 cells were scored for micronuclei, nuclear buds, binucleated cells, pyknotic cells, karyorrhectic cells, karyolytic cells, and cells with condensed chromatin. Group differences and trends were assessed across ordered HPV/cytology categories; genotype analyses used age-adjusted models and, among HPV-positive women, models adjusted for age and cytology. Women were classified as HPV-negative with negative for intraepithelial lesion or malignancy (HPV-/NILM, n = 69), HPV-positive with NILM (HPV+/NILM, n = 47), HPV-positive with atypical squamous cells of undetermined significance or low-grade squamous intraepithelial lesion (HPV+/ASCUS-LSIL, n = 49), and HPV-positive with high-grade squamous intraepithelial lesion or worse (HPV+/≥HSIL, n = 19). Significant positive trends across these ordered categories were observed for micronuclei (<i>p</i> = 0.044), nuclear buds (<i>p</i> = 0.029), and binucleated cells (<i>p</i> = 0.011). Among HPV-positive women, genotype differences adjusted for age and cytology were observed for nuclear buds (<i>p</i> = 0.046), karyolytic cells (<i>p</i> = 0.010), and condensed chromatin (<i>p</i> = 0.035). Overall, micronuclei, nuclear buds, and binucleated cells showed consistent patterns of genomic instability across HPV/cytology categories, while genotype-focused analyses also identified differences in selected genotoxic and cytotoxic endpoints. These findings support the potential value of MNcyt biomarkers as a complementary approach for risk assessment in HPV-related cervical disease.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"1-12"},"PeriodicalIF":1.4,"publicationDate":"2026-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149134768","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Quantification of microglial dynamic motility <i>in vivo</i>.","authors":"Ran Ran, Dong Wang, Jin Zhao, Linlin Hu, Yuan Zhao, Qingwu Yang, Sen Lin, Xiaofeng Cheng","doi":"10.1080/10520295.2026.2714773","DOIUrl":"10.1080/10520295.2026.2714773","url":null,"abstract":"<p><p>Motility is a fundamental property of microglial processes. Their dynamic movement is highly sensitive to alterations in the surrounding microenvironment and serves as an early indicator of pathological conditions in the central nervous system (CNS). Thus, comprehensive and accurate quantification of microglial process dynamics is critically needed. To address this, we developed a novel approach based on 3D alignment of original two-photon time-lapse images to correct misalignments caused by microglial migration or animal motion. This alignment facilitates subsequent colocalization analysis, allowing quantification of complete microglial structural dynamics at single-pixel resolution. Compared to conventional methods, our technique offers enhanced comprehensiveness and precision, enabling simultaneous measurement of process extension or retraction, microglial loss or regeneration, and clear visualization of temporal structural changes. Finally, we quantified the dynamic motility of microglia in abilateral common carotid artery occlusion/reperfusion (BCCAO/R) model using this method. We anticipate that this method will significantly improve the quantitative accuracy of dynamic motility assessments for microglia and other cell types <i>in vivo</i>.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"419-428"},"PeriodicalIF":1.4,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148807853","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Immunohistochemical and molecular insights into the anticoccidial, antioxidant, and antiapoptotic activities of <i>Coriandrum sativum</i> leaves extract.","authors":"Shaimaa M Kasem, Mohamed A Dkhil, Heba Ismaeil, Saeed El-Ashram, Andreas Meryk, Hend M Alharbi, Rewaida Abdel-Gaber, Mohammed Albeshr, Murad A Mubaraki, Nabila M Mira","doi":"10.1080/10520295.2026.2698596","DOIUrl":"10.1080/10520295.2026.2698596","url":null,"abstract":"<p><p>Coccidiosis is a significant enteric disease impacting economically important livestock, caused by a multidrug-resistant parasite. This research was performed to assess the immunohistochemical and molecular insights of <i>Coriandrum sativum</i> leaves extract (CSLE) against <i>Eimeria papillata</i> infection in murine models. CSLE subjected to phytochemical analysis. Thirty male Swiss albino mice were included: Group 1: Control; Group 2: Supplemented group receiving CSLE at 300 mg/kg B.W. (CSLE); Group 3: Infected with 1000 sporulated <i>E. papillata</i> oocysts; Group 4: Infected-treated with CSLE at 300 mg/kg B.W. (Infected + CSLE). Group 5: Infected-treated with amprolium at 120 mg/kg B.W. (Infected+Amp). Following 60 min of infection, groups 4 and 5 received daily treatment for five successive days with CSLE and Amp, respectively. Measurements of oocyst suppression and jejunal glutathione peroxidase (GPx) activity were conducted. Concentrations of Caspase-3 (Casp3) as well as jejunal nuclear factor kappa B (NFκB) and Casp-3 cells were measured immunohistochemically. Gene expression of apoptotic genes was assessed. The CSLE exhibited a substantial concentration of total phenolics and flavonoids, in addition to notable antioxidant activity (48.04 ± 2.98% DPPH scavenging). CSLE treatment exhibited significant oocyst suppression and minimized parasitic stages in the jejunum. CSLE reinstated jejunal GPx activity and mitigated the infection-induced elevation in Casp3 protein levels and its gene expression. Furthermore, CSLE modified apoptotic gene expressions, significantly downregulating BCL-2-associated protein x (BAX) and upregulating B-cell leukemia/lymphoma 2 (BCL2) following CSLE therapy. These data collectively underscore the significant anticoccidial, antioxidant, and antiapoptotic capabilities of CSLE, indicating its potential as a natural therapeutic treatment for <i>E. papillata</i> infection.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"387-397"},"PeriodicalIF":1.4,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148454440","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Turmeric as a counterstain in histology.","authors":"Carolina Wilson, Lindsey K Gregori, Sheila Criswell","doi":"10.1080/10520295.2026.2700352","DOIUrl":"10.1080/10520295.2026.2700352","url":null,"abstract":"<p><p>Histologic stains which identify specific tissue and cellular components are a staple of all diagnostic pathology laboratories. Histotechnology scientists are constantly attempting to find methods and reagents which are specific, simple to use, and cost effective. Turmeric (<i>Curcuma longa</i>) is a yellow spice readily available in grocery stores and is inexpensive. Previously, turmeric has been investigated as a natural histological stain, with most studies describing predominantly cytoplasmic staining and suggesting turmeric as a replacement for eosin. However, the present study supports the use of turmeric as a counterstain as staining localization is strongly dependent on the solvent, co-solute and procedure used for extraction and for staining. In contrast to several prior reports using alcoholic turmeric extracts, alcohol-based preparations in this study produced little to no appreciable tissue staining despite exhibiting intense coloration in solution. Instead, an aqueous extract with addition of aluminum was the most effective. A 0.5% turmeric solution prepared in aqueous 5% aluminum sulfate produced a bright yellow stain in histologic sections. While yellow chromatin against a colorless background provided limited contrast as a routine nuclear stain substitute, pairing turmeric with selected companion stains yielded favorable results. As a counterstain for alcian blue, Prussian blue, von Kossa, and rhodanine methods, turmeric provided excellent visual contrast. Turmeric is also markedly less expensive than nuclear fast red, supporting its potential as an economical alternative for selected histological applications.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"398-404"},"PeriodicalIF":1.4,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148454455","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Primary cell cultures versus immortalized cell lines: critical comparative insights into biological fidelity and translational applications.","authors":"Latifah Suali, Yvonne Chia, Christianus Rustin, Fui Fui Lem, Xin Jie Ching, Mila Atiqah Binti Mansur, Zachary Alden Henry, Mike K S Chan, Michelle B F Wong, Shing Yi Pan","doi":"10.1080/10520295.2026.2714776","DOIUrl":"10.1080/10520295.2026.2714776","url":null,"abstract":"<p><p><i>In vitro</i> cell culture models are fundamental to biomedical research, yet the choice between primary cell cultures and immortalized cell lines profoundly influences experimental outcomes. Immortalized lines offer convenience and unlimited proliferative capacity but frequently acquire genetic, epigenetic, and phenotypic alterations that compromise tissue-specific functions and overall biological fidelity. In contrast, primary cell cultures preserve native morphology, signaling pathways, and functional characteristics of their tissue of origin, providing a more physiologically relevant model. While previous reviews have outlined the general advantages of primary cells, few critically synthesize comparative data across cell types or evaluate strategies to mitigate inherent limitations such as finite lifespan and donor variability. This review critically examines the comparison between primary and immortalized cells across different cell types in terms of preserved tissue-specific characteristics, genomic and epigenetic fidelity, predictive value in drug testing and disease modeling, and emerging roles in personalized medicine. We further discuss innovative approaches, including conditional reprogramming, optimized donor selection, pooled-donor strategies, and standardized culture protocols, that enhance reproducibility and functional stability. By integrating comparative evidence with methodological advances, this review provides a critical framework for selecting appropriate <i>in vitro</i> models, highlighting the translational significance of primary cells in drug discovery, precision medicine, and biomedical research.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"429-455"},"PeriodicalIF":1.4,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148788022","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The effect of losartan nanoparticles vs losartan on TIMP1 and α-SMA immunohistochemical and gene expression in carbon tetrachloride-induced liver fibrosis.","authors":"Shrouk B Mohamed, Kamal G Botros, Salwa M Abo El-Khair, Mamdouh Eldesoqui, Wael Mohammed Elsaed, Sally Abdallah Mostafa, Yasir Hassan Elhassan, Islam Mohamed Mustafa Salama, Rasha Hamed Al-Serwi, Mosaab Salah El-Din El-Agawy","doi":"10.1080/10520295.2026.2703592","DOIUrl":"10.1080/10520295.2026.2703592","url":null,"abstract":"<p><p>Liver fibrosis is characterized by scar formation as a response to injury. Although many drugs have demonstrated antifibrotic efficacy in vitro and in animal models, effectively translating these into effective clinical therapies remains a challenge. Nanotechnology offers potential therapeutic approaches for treating liver fibrosis. This study aimed to assess and compare the antifibrotic effects of losartan and losartan nanoparticles (LP-NPs) on CCl₄-induced liver fibrosis in rats.Thirty-two adult male Sprague-Dawley rats were divided into four groups: a control group, a CCl₄-induced fibrosis group, a losartan-treated group, and an LP-NPs-treated group. Liver fibrosis was induced by CCl₄ injections over six weeks. Losartan or LP-NPs were orally administered for four weeks. At the conclusion of the study, rats were euthanized, and blood samples were collected for liver enzyme assessment. Liver specimens were subjected to histopathological examination (H&E, Sirius red staining), immunohistochemistry (α-SMA and TIMP1), and quantitative real-time PCR (qRT-PCR) for gene expression analysis of α-SMA and TIMP1. Morphometrical studies quantified collagen areas in Sirius red-stained sections and optical density of α-SMA and TIMP1 in immunohistochemically stained sections.The results showed that CCl₄ induced significantly elevated liver enzymes, severe distortion of hepatic architecture, marked steatosis and fibrosis. Additionally, there was an increase in collagen content, accompanied by strong positive immunostaining and upregulated gene expression of α-SMA and TIMP1. Both losartan and LP-NPs treatments significantly improved these parameters compared to the CCl₄-only group. Notably, LP-NPs demonstrated significantly greater improvements in liver enzyme levels, collagen percentage area, and the gene expression of α-SMA and TIMP1 compared to the standard losartan group. However, no significant difference was observed between the two treatments in α-SMA optical density.In conclusion, this investigation demonstrates that nanoformulation significantly enhanced losartan's antifibrotic efficacy. LP-NPs improved liver function and histological architecture, reduced collagen content after CCl₄ administration, and demonstrated greater therapeutic efficacy than conventional losartan.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"405-418"},"PeriodicalIF":1.4,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148788017","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Selective visualisation of aromatic and aliphatic suberin domains using combined Sudan Red-7B staining and UV autofluorescence.","authors":"Rachel Predeepa-Javahar","doi":"10.1080/10520295.2026.2720913","DOIUrl":"10.1080/10520295.2026.2720913","url":null,"abstract":"<p><p>This article reports a visualization method that combines Sudan red 7B staining with UV autofluorescence in rice root secondary cell walls. Sudan red 7B stains the aliphatic domains of suberin (SPAD), as well as intracellular lipids, whereas after UV excitation of unstained sections the aromatic domains of suberin (SPPD) emitted violet autofluorescence. However, when Sudan red 7B staining preceded UV excitation, the suberized cell walls fluoresced in varying colors from red to magenta, violet or bluish white. The optical phenomenon, Inner Filter Effect (IFE), exhibited this color variation as the differential solubility of Sudan Red 7B stain-SPAD suberin domain complex acted as an optical filter that shifted the violet autofluorescence from SPPD suberin domains to red. At the time of visualization, the red fluorescence spectrally mixed with violet autofluorescence such that the suberized cell walls fluoresced in varying colors. Using this visualization procedure, the intramolecular deposition and distribution pattern for SPAD and SPPD suberin domains in the outer apoplastic barrier can be effectively ascertained.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"456-461"},"PeriodicalIF":1.4,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148905179","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Histochemical staining optimization of a sperm chromatin dispersion test for the evaluation of DNA fragmentation in bovine spermatozoa.","authors":"Vasiliki Sapanidou, Konstantinos Petropoulos, Dimitrios Kalogiannis, Christos Balaskas","doi":"10.1080/10520295.2026.2737877","DOIUrl":"https://doi.org/10.1080/10520295.2026.2737877","url":null,"abstract":"<p><p>Sperm DNA integrity is a crucial determinant of fertility and early embryonic development. The Sperm Chromatin Dispersion (SCD) test provides a simple, practical, and cost-effective tool for assessing DNA fragmentation; however, the diagnostic reliability depends heavily on the staining technique used to visualize halos of dispersed chromatin. This study evaluated and optimized several commonly and widely used histochemical stains associated with a modified SCD protocol in bovine spermatozoa to improve accuracy and interpretation; hence providing new and useful data for scientists in the field. Frozen-thawed semen samples from five fertile bulls were processed using SCD procedures and stained with Toluidine Blue (TB), Giemsa (G), May Grünwald-Giemsa (MGG), Eosin Thiazine (ET), Feulgen & Rossenbeck (F&R), Hematoxylin & Eosin (H&E), Nuclear Fast Red (NFR), Eosin-Nigrosine (EN), and various modifications of the above stains. Most stains and their modifications (TB, G, ET, NFR, EN) failed to provide consistent or distinguishable halos, whilst specific modifications of MGG, F&R, and H&E produced reproducible halos. The MGG-modified protocol enabled halo visualization but introduced notable nonspecific background staining that reduced distinctiveness of halos. Specific modified protocols of F&R and H&E also visualized halos but reacted variably with agarose: F&R did not produce nonspecific background staining while H&E did. In conclusion, our results demonstrate that specific modifications of MGG, F&R, and H&E visualized halos and thus are suitable for SCD test.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"1-14"},"PeriodicalIF":1.4,"publicationDate":"2026-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149079531","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Pathologic and immunohistochemical findings in sea bass (Dicentrarchus labrax) naturally infected with <i>Aeromonas veronii biovar veronii</i>.","authors":"Emre Berke, Ayşe Nur Akkoç, Nihat Toplu","doi":"10.1080/10520295.2026.2726222","DOIUrl":"https://doi.org/10.1080/10520295.2026.2726222","url":null,"abstract":"<p><p>Tissue samples from European sea bass (<i>Dicentrarchus labrax</i>) naturally infected with Aeromonas veronii biovar veronii were subjected to bacteriological, molecular, pathological, and immunohistochemical analyses. The fish were sourced from aquaculture farms in the Aegean region of Turkiye. Clinical and postmortem examinations revealed granuloma formation as the most prominent lesion, primarily affecting the skin, spleen, kidneys, and liver. Sequencing of the 16S rRNA gene from all isolates showed 96-98% identity with <i>A. veronii</i> biovar <i>veronii</i> in the BLASTN 2.13.0+ database. Histopathological analysis demonstrated that granulomas originated from clusters of epithelioid macrophages and progressed through five distinct stages, culminating in caseous necrosis. Immunohistochemical labeling localized <i>A. veronii</i> antigens within necrotic regions and within the cytoplasm of macrophages in granulomatous lesions. These findings suggest that macrophages serve as the primary host cells for the bacterium and play a key role in its intra- and inter-tissue dissemination.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"1-11"},"PeriodicalIF":1.4,"publicationDate":"2026-09-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149015514","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comparison of manual counting with two rule-based image analysis programs, ImageJ and CellProfiler, for quantitative analysis of c-Fos immunohistochemical positivity in rat brainstem sections.","authors":"Manal Y Udwan, Ayman M Al-Qaaneh, Mimas H Al-Helalat, Ayah A Bashkami, Ezidin G Kaddumi","doi":"10.1080/10520295.2026.2731026","DOIUrl":"https://doi.org/10.1080/10520295.2026.2731026","url":null,"abstract":"<p><p>Quantitative and semiquantitative assessment of biomarker expression using immunohistochemistry (IHC) can be performed using several analytical approaches. Manual counting remains widely used but is inherently subjective and susceptible to observer variability. Rule-based image-analysis platforms may provide accessible alternatives, highlighting the need for direct method-comparison and agreement studies. c-Fos is a well-established marker of neuronal activation commonly quantified by counting IHC-positive cells. This study compared manual counting, ImageJ, and CellProfiler for quantifying c-Fos-positive cells in rat brainstem sections. Of 125 images initially evaluated, one was excluded as an extreme outlier, leaving 124 images for final analysis. Each image was independently analyzed by two trained observers using all three methods. ImageJ and CellProfiler were applied through their standard graphical interfaces and built-in functions using predefined user-guided threshold ranges, without additional plugins, scripts, machine-learning models, or custom code. Agreement was assessed using intraclass correlation coefficients (ICCs) and Bland-Altman analysis. No statistically significant difference in c-Fos-positive cell counts was observed among the three methods (Friedman test, P = 0.07). Median (IQR) counts were 6.75 (0.50-13.50) for manual counting, 6.25 (1.00-14.38) for ImageJ, and 6.00 (0.63-14.50) for CellProfiler. Inter-observer agreement was excellent for manual counting (ICC = 0.983), ImageJ (ICC = 0.964), and CellProfiler (ICC = 0.962). Inter-method agreement was also excellent (single-measures ICC = 0.971; 95% CI = 0.961-0.979). Bland-Altman analysis showed small mean biases of 0.33, 0.27, and -0.07 cells, with no significant proportional bias. Manual counting, ImageJ, and CellProfiler produced statistically comparable c-Fos-positive cell counts with excellent agreement. Under the predefined user-guided threshold ranges used, both software platforms closely matched manual counting; however, image-dependent threshold adjustment remains a potential source of observer variability.</p>","PeriodicalId":8970,"journal":{"name":"Biotechnic & Histochemistry","volume":" ","pages":"1-11"},"PeriodicalIF":1.4,"publicationDate":"2026-09-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148987929","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}