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Mpox, caused by viruses of the genus Orthopoxvirus, is an emerging threat to human and animal health. With increasing urbanization and more frequent interaction between humans and wild animals, the risk of Mpox transmission to humans has increased significantly. This review aims to examine in depth the epidemiology, pathogenesis, and diagnosis of Mpox, with a special focus on recent discoveries and advances in understanding the disease. Molecular mechanisms involved in viral replication will be examined, as well as risk factors associated with interspecific transmission and spread of the disease in human populations. Currently available diagnostic methods will also be discussed, with a critical analysis of their limitations and possible future directions for improving the accuracy and timeliness of diagnosis. Finally, this review will explore the public health implications associated with Mpox, emphasizing the importance of epidemiological surveillance, vaccination, and emergency preparedness to prevent and manage possible outbreaks. Understanding the epidemiology and control strategies for Mpox is critical to protecting the health of human and animal communities and mitigating the risk of interspecific transmission and spread of the disease. [ABSTRACT FROM AUTHOR]

Identification of biomarkers could help in assessing periodontal health status and monitoring treatment outcomes. Therefore, the aim of this cross-sectional study was to identify potential innovative salivary biomarkers for the diagnosis of periodontitis using an untargeted proteomic approach. Forty-five healthy non-smoker participants diagnosed as having periodontally healthy conditions (H), severe periodontitis (P), and healthy but reduced periodontium after active periodontal treatment (T) were consecutively enrolled (15 per each group) in the study. A higher number of spots were identified in the proteome of unstimulated whole saliva collected from H and T subjects compared with P group, mainly within the range of 8–40 kDa. Protein spots of interest were analysed by MALDI-TOF-MS, allowing the identification of cystatin SN (CST1) isoform, as confirmed by Western blot. CST1 was markedly expressed in the H group, while it was absent in most P samples (p < 0.001). Interestingly, a distinct CST1 expression was observed in saliva from T patients. CST1 was negatively correlated with the percentage of pathological sites (p < 0.001) and was effective in discriminating active periodontitis from healthy periodontal status (whether H or T). Therefore, salivary CST1 may be a promising non-invasive biomarker for periodontal disease diagnosis and monitoring. [ABSTRACT FROM AUTHOR]

Screening programs may be useful to reduce the rate of undetected glaucoma. Compass (CMP, CenterVue, Padova, Italy) Standard Suprathreshold strategy (SST) analyses the visual function at 52 retinal locations. A new Quick Suprathreshold strategy (QST) reduces the number of tested locations to 24. With both tests, the CMP also provides an image of the central retina and a detail of the optic nerve head. The aim of this paper is to measure the performances of SST and QST compared with clinical diagnosis. 63 consecutive healthy subjects and 60 consecutive patients with perimetric defects from glaucoma in both eyes were recruited. They received one test per eye (SST or QST in randomized order); results were classified into normal, suspect and abnormal according to a global index provided in the report and called SupraThreshold Response (STR). Agreement between clinical diagnosis and test output were calculated, and test time was also measured. The agreement with the clinical diagnosis was 95.7% for SST and 95.1% for QST. When two suspect tests were excluded, agreement for QST increased to 96.7%. Test duration was 164 ± 82 s for SST and 71 ± 41 s for QST (p < 0.0001). Such a difference was similar in both glaucoma patients (respectively 231 ± 65 s vs. 105 ± 33 s, p < 0.0001) and normal subjects (98 ± 17 and 39 ± 9 s, p < 0.0001). In conclusion, SST and QST showed similar, high agreement with clinical judgement. Morphological analysis is potentially helpful in further improving the clinical usefulness of suprathreshold tests. QST is characterized by a strong time saving compared with SST. [ABSTRACT FROM AUTHOR]

Thyroid cancer is the most common malignancy of the endocrine system, encompassing different entities with distinct histological features and clinical behavior. The diagnostic definition, therapeutic approach, and follow-up of thyroid cancers display some controversial aspects that represent unmet medical needs. Liquid biopsy is a non-invasive approach that detects and analyzes biological samples released from the tumor into the bloodstream. With the use of different technologies, tumor cells, free nucleic acids, and extracellular vesicles can be retrieved in the serum of cancer patients and valuable molecular information can be obtained. Recently, a growing body of evidence is accumulating concerning the use of liquid biopsy in thyroid cancer, as it can be exploited to define a patient's diagnosis, estimate their prognosis, and monitor tumor recurrence or treatment response. Indeed, liquid biopsy can be a valuable tool to overcome the limits of conventional management of thyroid malignancies. In this review, we summarize currently available data about liquid biopsy in differentiated, poorly differentiated/anaplastic, and medullary thyroid cancer, focusing on circulating tumor cells, circulating free nucleic acids, and extracellular vesicles. [ABSTRACT FROM AUTHOR]

Celiac disease (CD) is diagnosed by a combination of specific serology and typical duodenal lesions. The histological confirmation of CD, mandatory in the majority of patients with suspected CD, is based on invasive and poorly tolerated procedures, such as upper gastrointestinal endoscopy. In this study we propose an alternative and non-invasive methodology able to confirm the diagnosis of CD based on the analysis of serum samples using the Raman spectroscopy technique. Three different bands centered at 1650, 1450 and 1003 cm−1 have been considered and the A1450/A1003 and A1650/A1003 ratios have been computed to discriminate between CD and non-CD subjects. The reliability of the methodology was validated by statistical analysis using receiver operating characteristic (ROC) curves. The Youden index was also determined to obtain optimal cut-off points. The obtained results highlighted that the proposed methodology was able to distinguish between CD and non-CD subjects with 98% accuracy. The optimal cut-off points revealed, for both the A1450/A1003 and A1650/A1003 ratios, high values of sensitivity and specificity (>95.0% and >92.0% respectively), confirming that Raman spectroscopy may be considered a valid alternative to duodenal biopsy and demonstrates spectral changes in the secondary structures of the protein network. [ABSTRACT FROM AUTHOR]

Simple Summary: Oral squamous cell carcinoma (OSCC) accounts for 90–95% of malignant tumors of the lip and oral cavity and is associated with high mortality in the advanced stages. Early diagnosis is a challenge for oral pathologists and dentists, due to the ambiguous appearance of early OSCC, which is often misdiagnosed, mistreated, and associated with diagnostic delay. The gold standards for OSCC diagnosis are biopsy and histopathological assessment, but these procedures are invasive and time-consuming. Adjunctive noninvasive techniques allow the definition of the malignant features of a suspicious lesion in real time and noninvasively, thus improving the diagnostic procedure. The present review aimed to focus on some of the main promising noninvasive imaging techniques, to highlight their perspective adoption in a three-step diagnosis, which is idealistically faster and better, as well as enables the patient's compliance. Oral squamous cell carcinoma (OSCC) is the most prevalent form of cancer of lips and oral cavity, and its diagnostic delay, caused by misdiagnosis at the early stages, is responsible for high mortality ratios. Biopsy and histopathological assessment are the gold standards for OSCC diagnosis, but they are time-consuming, invasive, and do not always enable the patient's compliance, mainly in cases of follow-up with the need for more biopsies. The use of adjunctive noninvasive imaging techniques improves the diagnostic approach, making it faster and better accepted by patients. The present review aims to focus on the most consolidated diagnostic techniques, such as vital staining and tissue autofluorescence, and to report the potential role of some of the most promising innovative techniques, such as narrow-band imaging, high-frequency ultrasounds, optical coherence tomography, and in vivo confocal microscopy. According to their contribution to OSCC diagnosis, an ideal three-step diagnostic procedure is proposed, to make the diagnostic path faster, better, and more accurate. [ABSTRACT FROM AUTHOR]

Pattern analysis of the salivary metabolic profile has been proven accurate in discriminating between generalized periodontitis (GP) patients and healthy individuals (HI), as this disease modifies the salivary concentrations of specific metabolites. Due to the scarcity of data from previous studies, this study aimed to evaluate if non-surgical periodontal therapy (NST) could affect the metabolomic profile in GP patients' saliva and if it compares to that of HI. Unstimulated salivary samples were collected from 11 HI and 12 GP patients before and 3 months after NST. Nuclear Magnetic Resonance (NMR) spectroscopy, followed by a supervised multivariate statistical approach on entire saliva spectra and partial least square (PLS) discriminant analysis, were performed to obtain metabolic profiles. In the GP group, periodontal treatment improved all clinical parameters, but not all the diseased sites were eradicated. PLS revealed an accuracy of 100% in distinguishing between metabolic profiles of GP patients before and after NST. Orthogonal projection to latent structure was able to discriminate between the three groups of subjects with an accuracy of 85.6%. However, the post-NST metabolic profile of GP patients could not be completely assimilated to that of HI. Although NST may produce significant changes in the metabolic profile, GP patients maintained a distinctive fingerprint compared to HI. [ABSTRACT FROM AUTHOR]

This systematic review describes the several methods to diagnose and measure the severity of small fiber neuropathies and aims to guide the physician to define all the diagnostic approaches for adopting the best strategies described in the current literature. The search was conducted in PubMed, EMBASE, Cochrane Library and Web of Science. Two reviewers independently reviewed and came to consensus on which articles met inclusion/exclusion criteria. The authors excluded all the duplicates, animals' studies, and included the English articles in which the diagnostic measures were finalized to assess the effectiveness of rehabilitation and pharmacologic treatment of patients with small fiber neuropathies. The search identified a total of 975 articles with the keywords "small fiber neuropathy" AND "rehabilitation" OR "therapy" OR "treatment". Seventy-eight selected full-text were analyzed by the reviewers. Forty-one publications met the inclusion criteria and were included in the systematic review. Despite the range of diagnostic tools for the assessment of small fiber neuropathy, other robust trials are needed. In addition, always different diagnostic approaches are used, a unique protocol could be important for the clinicians. More research is needed to build evidence for the best diagnostic methodologies and to delineate a definitive diagnostic protocol. [ABSTRACT FROM AUTHOR]

Gadolinium-based contrast agents (GBCAs) have helped to improve the role of magnetic resonance imaging (MRI) for the diagnosis and treatment of diseases. There are currently nine different commercially available gadolinium-based contrast agents (GBCAs) that can be used for body MRI cases, and which are classifiable according to their structures (cyclic or linear) or biodistribution (extracellular-space agents, target/specific-agents, and blood-pool agents). The aim of this review is to illustrate the commercially available MRI contrast agents, their effect on imaging, and adverse reaction on the body, with the goal to lead to their proper selection in different clinical contexts. When we have to choose between the different GBCAs, we have to consider several factors: (1) safety and clinical impact; (2) biodistribution and diagnostic application; (3) higher relaxivity and better lesion detection; (4) higher stability and lower tissue deposit; (5) gadolinium dose/concentration and lower volume injection; (6) pulse sequences and protocol optimization; (7) higher contrast-to-noise ratio at 3.0 T than at 1.5 T. Knowing the patient's clinical information, the relevant GBCAs properties and their effect on body MRI sequences are the key features to perform efficient and high-quality MRI examination. [ABSTRACT FROM AUTHOR]

Magnetic resonance imaging (MRI) is the gold standard for glioblastoma (GBM) patient evaluation. Additional non-invasive diagnostic modalities are needed. GBM is heavily infiltrated with tumor-associated macrophages (TAMs) that can be found in peripheral blood. FKBP51s supports alternative-macrophage polarization. Herein, we assessed FKBP51s expression in circulating monocytes from 14 GBM patients. The M2 monocyte phenotype was investigated by qPCR and flow cytometry using antibodies against PD-L1, CD163, FKBP51s, and CD14. MRI assessed morphologic features of the tumors that were aligned to flow cytometry data. PD-L1 expression on circulating monocytes correlated with MRI tumor necrosis score. A wider expansion in circulating CD163/monocytes was measured. These monocytes resulted in a dramatic decrease in patients with an MRI diagnosis of complete but not partial surgical removal of the tumor. Importantly, in patients with residual tumor, most of the peripheral monocytes that in the preoperative stage were CD163/FKBP51s− had turned into CD163/FKBP51s+. After Stupp therapy, CD163/FKBP51s+ monocytes were almost absent in a case of pseudoprogression, while two patients with stable or true disease progression showed sustained levels in such circulating monocytes. Our work provides preliminary but meaningful and novel results that deserve to be confirmed in a larger patient cohort, in support of potential usefulness in GBM monitoring of CD163/FKBP51s/CD14 immunophenotype in adjunct to MRI. [ABSTRACT FROM AUTHOR]

Simple Summary: This manuscript aims to address the diagnostic challenges of mediastinal bulky lymphomas with the baseline value of 18F-FDG PET/CT metabolic, volumetric and texture parameters, also relying on machine learning techniques, in patients with grey zone lymphoma, primary diffuse large B-cell lymphoma of the mediastinum and classical Hodgkin lymphoma. Different types of histology demonstrated several baseline 18F-FDG PET/CT radiomics parameters that were significantly different from one another, suggesting the possibility of identifying potential histological heterogeneity and aggressive transformation. Moreover, using radiomics-based imaging biomarkers, machine learning techniques offer a solution for separating not completely disjoint histological types. To date, the gold standard for diagnosis is biopsy, but machine learning methods could be combined with radiomics to build a histological representation of mediastinal bulky masses that is able to successfully identify different types of lymphomas. Finally, this preliminary study supports the potential of metabolic texture analyses as future imaging biomarkers, with a growing role in clinical diagnosis. Background: This study tested the diagnostic value of 18F-FDG PET/CT (FDG-PET) volumetric and texture parameters in the histological differentiation of mediastinal bulky disease due to classical Hodgkin lymphoma (cHL), primary mediastinal B-cell lymphoma (PMBCL) and grey zone lymphoma (GZL), using machine learning techniques. Methods: We reviewed 80 cHL, 29 PMBCL and 8 GZL adult patients with mediastinal bulky disease and histopathological diagnoses who underwent FDG-PET pre-treatment. Volumetric and radiomic parameters were measured using FDG-PET both for bulky lesions (BL) and for all lesions (AL) using LIFEx software (threshold SUV ≥ 2.5). Binary and multiclass classifications were performed with various machine learning techniques fed by a relevant subset of radiomic features. Results: The analysis showed significant differences between the lymphoma groups in terms of SUVmax, SUVmean, MTV, TLG and several textural features of both first- and second-order grey level. Among machine learning classifiers, the tree-based ensembles achieved the best performance both for binary and multiclass classifications in histological differentiation. Conclusions: Our results support the value of metabolic heterogeneity as an imaging biomarker, and the use of radiomic features for early characterization of mediastinal bulky lymphoma. [ABSTRACT FROM AUTHOR]

Purpose: This study aims to explore and characterize healthy eye microbiota. Methods: Healthy subjects older than 18 years were selected for this descriptive cross-sectional study. Samples were collected with an eSwab with 1 mL of Liquid Amies Medium (Copan Brescia, Italy). Following DNA extraction, libraries preparation, and amplification, PCR products were purified and end-repaired for barcode ligation. Libraries were pooled to a final concentration of 26 pM. Template preparation was performed with Ion Chef according to Ion 510, Ion 520, and Ion 530 Kit-Chef protocol. Sequencing of the amplicon libraries was carried out on a 520 or 530 chip using the Ion Torrent S5 system (Thermo Fisher; Waltham, MA, USA). Raw reads were analyzed with GAIA (v 2.02). Results: Healthy eye microbiota is a low-diversity microbiome. The vast majority of the 137 analyzed samples were highly enriched with Staphylococcus, whereas only in a few of them, other genera such as Bacillus, Pseudomonas, and Corynebacterium predominate. We found an average of 88 genera with an average Shannon index of 0.65. Conclusion: We identified nine different ECSTs. A better understanding of healthy eye microbiota has the potential to improve disease diagnosis and personalized regimens to promote health. [ABSTRACT FROM AUTHOR]

Diagnosis of type I hypersensitivity reactions (IgE-mediated reactions) to penicillins is based on clinical history, skin tests (STs), and drug provocation tests (DPTs). Among in vitro complementary tests, the fluoro-enzyme immunoassay (FEIA) ImmunoCAP® (Thermo-Fisher, Waltham, MA, USA) is the most widely used commercial method for detecting drug-specific IgE (sIgE). In this study, we aimed to analyze the utility of ImmunoCAP® for detecting sIgE to penicillin G (PG) and amoxicillin (AX) in patients with confirmed penicillin allergy. The study includes 139 and 250 patients evaluated in Spain and Italy, respectively. All had experienced type I hypersensitivity reactions to penicillins confirmed by positive STs. Additionally, selective or cross-reactive reactions were confirmed by DPTs in a subgroup of patients for further analysis. Positive ImmunoCAP® results were 39.6% for PG and/or AX in Spanish subjects and 52.4% in Italian subjects. When only PG or AX sIgE where analyzed, the percentages were 15.1% and 30.4%, respectively, in Spanish patients; and 38.9% and 46% in Italian ones. The analysis of positive STs showed a statistically significant higher percentage of positive STs to PG determinants in Italian patients. False-positive results to PG (16%) were detected in selective AX patients with confirmed PG tolerance. Low and variable sensitivity values observed in a well-defined population with confirmed allergy diagnosis, as well as false-positive results to PG, suggest that ImmunoCAP® is a diagnostic tool with relevant limitations in the evaluation of subjects with type I hypersensitivity reactions to penicillins. [ABSTRACT FROM AUTHOR]

Background: Pneumoperitoneum is defined by the presence of free air in the abdominal cavity; gastrointestinal perforation is an important cause of this pathological condition. In emergency situations, radiology is considered vital in the early detection and identification of the site and cause of the perforation, which is critical for proper surgical planning. Aim: The aim of our study was to evaluate a new diagnostic US tool, based on the US contrast-specific software generally used during contrast-enhanced US examination (CEUS), without the administration of sonographic contrast media, and to describe the specific imaging features in the detection of free intra-peritoneal air. Subjects and Methods: One hundred and fifty-seven consecutive and hemodynamically stable patients, who arrived in our E.D. with an acute abdomen between April 2018 and October 2019, underwent US and CT examination, performed by three radiologists (with 5, 5, and 25 years of experience). The US was performed first and divided into two steps, using B-mode US and both B-mode and contrast-specific software US, with no contrast media administration. All the patients underwent CT examination. Results: In 32 out of 157 patients, the surgery confirmed GI perforation. CT correctly detected 31 out of 32 patients; the contrast-specific software US identified 30 perforated patients. CT reached a sensitivity value of 97% and specificity value of 100%; contrast-specific software US demonstrated higher values than B-mode US in sensitivity (93% vs. 70%, respectively) and specificity (98% vs. 88%, respectively). Conclusion: the use of contrast-specific software in emergencies improves image quality, and reaches higher levels of sensitivity and specificity with no time delay compared to standard US examination, helping radiologists expedite diagnoses. [ABSTRACT FROM AUTHOR]

Among the tools useful for the management of neuropathic pain, ultrasound presents several advantages, shown by the literature. We assessed the scientific production about neuropathic pain and ultrasound from different points of view: general topics, journal categories, geographical origin and lexical analysis. We searched papers on PubMed using the Medical Subject Headings "neuropathic pain" AND "ultrasound". We collected data about the journals where the papers were published, the country of the affiliation of the first author. For the lexical analysis, we evaluated the presence of selected words in the papers, and we built a graph representing the connections among words and papers. The papers were focused on the use of ultrasound as a diagnostic tool and guide for the therapy, assessing its application in different diseases such as Morton's neuroma and piriformis syndrome. The most represented journal category was anesthesia while the most common country the United States of America. The lexical analysis confirmed the importance of ultrasound for diagnosis of specific disease and treatment of pain. The described approaches provide a multiperspective evaluation of the literature and may support the interpretation of the information contained by the papers. [ABSTRACT FROM AUTHOR]

Systemic lupus erythematosus (SLE) is a clinically and genetically heterogeneous autoimmune disease. The etiology of lupus and the contribution of genetic, environmental, infectious and hormonal factors to this phenotype have yet to be elucidated. The most straightforward approach to unravel the molecular pathogenesis of lupus may rely on studies of patients who present with early-onset severe phenotypes. Typically, they have at least one of the following clinical features: childhood onset of severe disease (<5 years), parental consanguinity, and presence of family history for autoimmune diseases in a first-degree relative. These patients account for a small proportion of patients with lupus but they inform considerable knowledge about cellular pathways contributing to this inflammatory phenotype. In recent years with the aid of new sequencing technologies, novel or rare pathogenic variants have been reported in over 30 genes predisposing to SLE and SLE-like diseases. Future studies will likely discover many more genes with private variants associated to lupus-like phenotypes. In addition, genome-wide association studies (GWAS) have identified a number of common alleles (SNPs), which increase the risk of developing lupus in adult age. Discovery of a possible shared immune pathway in SLE patients, either with rare or common variants, can provide important clues to better understand this complex disorder, it's prognosis and can help guide new therapeutic approaches. The aim of this review is to summarize the current knowledge of the clinical presentation, genetic diagnosis and mechanisms of disease in patents with lupus and lupus-related phenotypes. [ABSTRACT FROM AUTHOR]

Macular degenerations represent leading causes of central blindness or low vision in developed countries. Most of these severe visual disabilities are due to age-related macular degeneration (AMD) and pathologic myopia (PM), both of which are frequently complicated by subfoveal choroidal neovascularization (CNV). Photodynamic therapy with verteporfin (PDT-V) is still employed for CNV treatment in selected cases or in combined regimen. In Caucasian patients, the common polymorphism G185T of factor XIII-A gene (FXIII-A-G185T; rs5985) has been described as predictor of poor angiographic CNV responsiveness to PDT-V. Nevertheless, the prognostic implications of this pharmacogenetic determinant on long-term visual outcome after a PDT-V regimen have not been evaluated. We retrospectively selected Caucasian patients presenting with treatment-naive CNV and receiving standardized PDT-V protocol for two years. The study population included patients affected by subfoveal CNV secondary to AMD or PM. We assessed the correlations between the polymorphic allele T of FXIII-A-G185T and: (1) total number of photodynamic treatments; and (2) change in visual acuity from baseline to the end of the follow-up period. Considering a total study population of 412 patients with neovascular AMD or PM, the carriers of 185 T-allele of FXIII-A (GT or TT genotype) received a higher number of photodynamic treatments than patients without it (GG wild-type genotype) (p < 0.01; mean number of PDT-V: 5.51 vs. 3.76, respectively). Moreover, patients with 185 T-allele of FXIII-A had a more marked worsening of visual acuity at 24 months than those with the GG-185 wild genotype (p < 0.01; mean difference in logMAR visual acuity: 0.22 vs. 0.08, respectively). The present findings show that the G185T polymorphism of the FXIII-A gene is associated with significant differences in the long-term therapeutic outcomes of patients treated with standardized PDT-V protocol. The comprehensive appraisal of both anti-thrombophilic effects due to FXIII-A G185T variant and photo-thrombotic action of PDT-V toward CNV provides several clues about the rationale of this intriguing pharmacogenetic correlation. Further investigations are warranted to outline the appropriate paradigm for guiding PDT-V utilization in the course of the combined therapeutic protocol for neovascular macular degeneration. [ABSTRACT FROM AUTHOR]

Trauma is one of the most common causes of death or permanent disability in young people, so a timely diagnostic approach is crucial. In polytrauma patients, CEUS (contrast enhanced ultrasound) has been shown to be more sensitive than US (ultrasound) for the detection of solid organ injuries, improving the identification and grading of traumatic abdominal lesions with levels of sensitivity and specificity similar to those seen with MDCT (multidetector tomography). CEUS is recommended for the diagnostic evaluation of hemodynamically stable patients with isolated blunt moderate-energy abdominal traumas and the diagnostic follow-up of conservatively managed abdominal traumas. In this pictorial review, we illustrate the advantages and disadvantages of CEUS and the procedure details with tips and tricks during the investigation of blunt moderate-energy abdominal trauma as well as during follow-up in non-operative management. [ABSTRACT FROM AUTHOR]

Yardlong bean (Vigna unguiculata subsp. sesquipedalis) plants with virus-like systemic mottling and leaf distortion were observed in both experimental and commercial fields in Aragua State, Venezuela. Symptomatic leaves were shown to contain carlavirus-like particles. RT-PCR analysis with carlavirus-specific primers was positive in all tested samples. Nucleotide sequences of the obtained amplicons showed 84%-74% similarity to corresponding sequences of Cowpea mild mottle virus (CPMMV) isolates deposited in the GenBank database. This is the first report of CPMMV in Venezuela and is thought to be the first report of CPMMV infecting yardlong bean. [ABSTRACT FROM AUTHOR]

Simple Summary: Pediatric solid tumors are a heterogenous group of diseases that comprise ≈ 40% of all pediatric cancers, early diagnosis being key for improved survival. Here we designed, tested, and validated a single eight-color tube for the diagnostic screening of pediatric cancer—solid tumor orientation tube (STOT)—based on multiparameter flow cytometry vs. conventional diagnostic procedures. Prospective clinical validation of STOT in 149 samples (63 tumor mass, 38 bone marrow, 30 lymph node, and 18 body fluid samples) screened for pediatric cancer, apart from 26 blood specimens that were excluded from analysis, showed concordant results with the final WHO/ICCC-3 diagnosis in 138/149 cases (92.6%). This included correct diagnostic orientation by STOT in 43/44 (98%) malignant and 4/4 (100%) benign non-hematopoietic tumors, together with 28/38 (74%) leukemia/lymphoma cases. The only recurrently missed diagnosis was Hodgkin lymphoma (0/8), which would require additional markers. These results support the use of STOT as a complementary tool for fast and accurate diagnostic screening, orientation, and classification of pediatric cancer in suspicious patients. Early diagnosis of pediatric cancer is key for adequate patient management and improved outcome. Although multiparameter flow cytometry (MFC) has proven of great utility in the diagnosis and classification of hematologic malignancies, its application to non-hematopoietic pediatric tumors remains limited. Here we designed and prospectively validated a new single eight-color antibody combination—solid tumor orientation tube, STOT—for diagnostic screening of pediatric cancer by MFC. A total of 476 samples (139 tumor mass, 138 bone marrow, 86 lymph node, 58 peripheral blood, and 55 other body fluid samples) from 296 patients with diagnostic suspicion of pediatric cancer were analyzed by MFC vs. conventional diagnostic procedures. STOT was designed after several design–test–evaluate–redesign cycles based on a large panel of monoclonal antibody combinations tested on 301 samples. In its final version, STOT consists of a single 8-color/12-marker antibody combination (CD99-CD8/numyogenin/CD4-EpCAM/CD56/GD2/smCD3-CD19/cyCD3-CD271/CD45). Prospective validation of STOT in 149 samples showed concordant results with the patient WHO/ICCC-3 diagnosis in 138/149 cases (92.6%). These included: 63/63 (100%) reactive/disease-free samples, 43/44 (98%) malignant and 4/4 (100%) benign non-hematopoietic tumors together with 28/38 (74%) leukemia/lymphoma cases; the only exception was Hodgkin lymphoma that required additional markers to be stained. In addition, STOT allowed accurate discrimination among the four most common subtypes of malignant CD45− CD56++ non-hematopoietic solid tumors: 13/13 (GD2++ numyogenin− CD271−/+ nuMyoD1− CD99− EpCAM−) neuroblastoma samples, 5/5 (GD2− numyogenin++ CD271++ nuMyoD1++ CD99−/+ EpCAM−) rhabdomyosarcomas, 2/2 (GD2−/+ numyogenin− CD271+ nuMyoD1− CD99+ EpCAM−) Ewing sarcoma family of tumors, and 7/7 (GD2− numyogenin− CD271+ nuMyoD1− CD99− EpCAM+) Wilms tumors. In summary, here we designed and validated a new standardized antibody combination and MFC assay for diagnostic screening of pediatric solid tumors that might contribute to fast and accurate diagnostic orientation and classification of pediatric cancer in routine clinical practice. [ABSTRACT FROM AUTHOR]