SpaceANOVA: Spatial Co-occurrence Analysis of Cell Types in Multiplex Imaging Data Using Point Process and Functional ANOVA.

Multiplex imaging platforms have enabled the identification of the spatial organization of different types of cells in complex tissue or the tumor microenvironment. Exploring the potential variations in the spatial co-occurrence or colocalization of different cell types across distinct tissue or disease classes can provide significant pathological insights, paving the way for intervention strategies. However, the existing methods in this context either rely on stringent statistical assumptions or suffer from a lack of generalizability. We present a highly powerful method to study differential spatial co-occurrence of cell types across multiple tissue or disease groups, based on the theories of the Poisson point process and functional analysis of variance. Notably, the method accommodates multiple images per subject and addresses the problem of missing tissue regions, commonly encountered due to data-collection complexities. We demonstrate the superior statistical power and robustness of the method in comparison with existing approaches through realistic simulation studies. Furthermore, we apply the method to three real data sets on different diseases collected using different imaging platforms. In particular, one of these data sets reveals novel insights into the spatial characteristics of various types of colorectal adenoma.

Vegan grade medium component screening and concentration optimization for the fermentation of the probiotic strain Lactobacillus paracasei IMC 502® using Design of Experiments.

Lactobacillus paracasei IMC502® is a commercially successful probiotic strain. However, there are no reports that investigate growth medium composition in relation to improved biomass production for this strain. The major outcome of the present study is the design and optimization of a growth medium based on vegan components to be used in the cultivation of Lactobacillus paracasei IMC502®, by using Design of Experiments. Besides comparing different carbon sources, the use of plant-based peptones as nitrogen sources was considered. In particular, the use of guar peptone as the main nitrogen source, in the optimization of fermentation media for the production of probiotics, could replace other plant peptones (e.g. potato, rice, wheat, and soy) which are part of the human diet, thereby avoiding an increase in product and process prices. A model with R2 and adjusted R2 values higher than 95% was obtained. Model accuracy was equal to 94.11%. The vegan-optimized culture medium described in this study increased biomass production by about 65% compared to growth on De Man-Rogosa-Sharpe (MRS) medium. Moreover, this approach showed that most of the salts and trace elements generally present in MRS are not affecting biomass production, thus a simplified medium preparation can be proposed with higher probiotic biomass yield and titer. The possibility to obtain viable lactic acid bacteria at high density from vegetable derived nutrients will be of great interest to specific consumer communities, opening the way to follow this approach with other probiotics of impact for human health.

Dietary Sugar and Anthropometrics among Young Children in the Guelph Family Health Study: Longitudinal Associations.

Purpose: Our understanding of the influence of sugar intake on anthropometrics among young children is limited. Most existing research is cross-sectional and has focused on sugar-sweetened beverages. The study objective was to investigate longitudinal associations between young children's total, free, and added sugar intake from all food sources at baseline with anthropometric measures at baseline and 18 months.Methods: The Guelph Family Health Study (GFHS) is an ongoing randomized controlled trial and a family-based health promotion study. Food records and anthropometric data were collected at baseline (n = 109, 55 males; 3.7 ± 1.1 y, mean ± SD) and 18 months (n = 109, 55 males; 5.1 ± 1.1 y) of the GFHS pilots. Associations between sugar intakes and anthropometrics were estimated using linear regression models with generalized estimating equations adjusted for age, sex, household income, and intervention status.Results: Total sugar intake was inversely associated with body weight at 18 months (P = 0.01). There was no effect of time on any other associations between total, free, and added sugar intakes and anthropometrics.Conclusions: Early life dietary sugar intakes may not relate to anthropometric measures in the short term. Further investigation into potential associations between dietary sugar intakes and anthropometric variables over longer time periods is warranted.

Molecular architecture of proliferative lupus nephritis as elucidated using 50-plex imaging mass cytometry proteomics.

Due to unique advantages that allow high-dimensional tissue profiling, we postulated imaging mass cytometry (IMC) may shed novel insights on the molecular makeup of proliferative lupus nephritis (LN). This study interrogates the spatial expression profiles of 50 target proteins in LN and control kidneys. Proliferative LN glomeruli are marked by podocyte loss with immune infiltration dominated by CD45RO+, HLA-DR+ memory CD4 and CD8 T-cells, and CD163+ macrophages, with similar changes in tubulointerstitial regions. Macrophages are the predominant HLA-DR expressing antigen presenting cells with little expression elsewhere, while macrophages and T-cells predominate cellular crescents. End-stage sclerotic glomeruli are encircled by an acellular fibro-epithelial Bowman's space surrounded by immune infiltrates, all enmeshed in fibronectin. Proliferative LN also shows signs indicative of epithelial to mesenchymal plasticity of tubular cells and parietal epithelial cells. IMC enabled proteomics is a powerful tool to delineate the spatial architecture of LN at the protein level.

Trends and Challenges in AIoT/IIoT/IoT Implementation.

For the next coming years, metaverse, digital twin and autonomous vehicle applications are the leading technologies for many complex applications hitherto inaccessible such as health and life sciences, smart home, smart agriculture, smart city, smart car and logistics, Industry 4.0, entertainment (video game) and social media applications, due to recent tremendous developments in process modeling, supercomputing, cloud data analytics (deep learning, etc.), communication network and AIoT/IIoT/IoT technologies. AIoT/IIoT/IoT is a crucial research field because it provides the essential data to fuel metaverse, digital twin, real-time Industry 4.0 and autonomous vehicle applications. However, the science of AIoT is inherently multidisciplinary, and therefore, it is difficult for readers to understand its evolution and impacts. Our main contribution in this article is to analyze and highlight the trends and challenges of the AIoT technology ecosystem including core hardware (MCU, MEMS/NEMS sensors and wireless access medium), core software (operating system and protocol communication stack) and middleware (deep learning on a microcontroller: TinyML). Two low-powered AI technologies emerge: TinyML and neuromorphic computing, but only one AIoT/IIoT/IoT device implementation using TinyML dedicated to strawberry disease detection as a case study. So far, despite the very rapid progress of AIoT/IIoT/IoT technologies, several challenges remain to be overcome such as safety, security, latency, interoperability and reliability of sensor data, which are essential characteristics to meet the requirements of metaverse, digital twin, autonomous vehicle and Industry 4.0. applications.

Novel Synthesis of IMC-48 and Affinity Evaluation with Different i-Motif DNA Sequences.

During the last decade, the evidence for the biological relevance of i-motif DNA (i-DNA) has been accumulated. However, relatively few molecules were reported to interact with i-DNA, and a controversy concerning their binding mode, affinity, and selectivity persists in the literature. In this context, the cholestane derivative IMC-48 has been reported to modulate bcl-2 gene expression by stabilizing an i-motif structure in its promoter. In the present contribution, we report on a novel, more straightforward, synthesis of IMC-48 requiring fewer steps compared to the previous approach. Furthermore, the interaction of IMC-48 with four different i-motif DNA sequences was thoroughly investigated by bio-layer interferometry (BLI) and circular dichroism (CD) spectroscopy. Surprisingly, our results show that IMC-48 is a very weak ligand of i-DNA as no quantifiable interaction or significant stabilization of i-motif structures could be observed, stimulating a quest for an alternative mechanism of its biological activity.

Lactobacillus paracasei IMC 502 ameliorates type 2 diabetes by mediating gut microbiota-SCFA-hormone/inflammation pathway in mice.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is a complex and prevalent metabolic disease that seriously threatens human health. Numerous studies have shown that probiotics as dietary supplements have the potential to prevent and treat T2DM. However, the ability of various strains to improve diabetes symptoms and corresponding mechanisms are different. Thus, mechanistic investigation is required to validate the pharmacology of each probiotic strain for T2DM treatment. Lactobacillus paracasei IMC 502 was originally isolated from Italian elderly human feces and its probiotic attributes have been demonstrated. Here, the antidiabetic pharmacodynamics of L. paracasei IMC 502 on T2DM mice was explored. RESULTS: Lactobacillus paracasei IMC 502 significantly decreased blood glucose, HbA1c and lipid levels, improved insulin resistance and glucose intolerance, regulated the mRNA/protein expression of key hepatic enzymes associated with gluconeogenesis, de novo lipogenesis and PI3K/Akt pathway, and repaired pancreatic and hepatic tissue damage. This probiotic conferred beneficial outcomes in the gut microbiome of diabetic mice, which induced transformation of short-chain fatty acids (SCFAs) and further enhanced the secretion of downstream hormones, and ultimately ameliorated the inflammatory response. CONCLUSION: Lactobacillus paracasei IMC 502 prevents and alleviates T2DM by mediating the gut microbiota-SCFA-hormone/inflammation pathway. © 2022 Society of Chemical Industry.

Ability of Anthropometric Measurements to Predict Metabolic Health among Patients in Alberta: A Cross-sectional Study in Primary Care.

Purpose: This study compared anthropometric and body fat percent (BF%) equations in relation to measures of metabolic health.Methods: BF% calculations (Bergman, Fels, and Woolcott) and anthropometric measurements were used to determine obesity among a sample of patients attending primary care in Alberta, Canada. Anthropometric variables included body mass index (BMI), waist circumference, waist:hip ratio, waist:height ratio, and calculated BF%. Metabolic Z-score was computed as the average of the individual Z-scores of triglycerides, total cholesterol, and fasting glucose and the number of standard deviations from the sample mean.Results: Five hundred and fourteen individuals were included (41.2% male, age: 53 ± 16y, BMI: 27.4 ± 5.7 kg/m2). BMI ≥ 30 kg/m2 detected the smallest number of participants (n = 137) as having obesity, while Woolcott BF% equation categorized the largest number of participants as having obesity (n = 369). No anthropometric or BF% calculation predicted metabolic Z-score in males (all p ≥ 0.05). In females, age-adjusted waist:height ratio had the highest prediction power (R2 = 0.204, p < 0.001), followed by age-adjusted waist circumference (R2 = 0.200, p < 0.001) and age-adjusted BMI (R2 = 0.178, p < 0.001).Conclusions: This study did not find evidence that BF% equations more strongly predicted metabolic Z-scores than other anthropometric values. In fact, all anthropometric and BF% variables were weakly related to metabolic health parameters, with apparent sex differences.

TITAN: An end-to-end data analysis environment for the Hyperion™ imaging system.

Imaging Mass Cytometry (IMC) is a powerful high-throughput technique enabling resolution of up to 37 markers in a single fixed tissue section while also preserving in situ spatial relationships. Currently, IMC processing and analysis necessitates the use of multiple different software, labour-intensive pipeline development, different operating systems and knowledge of bioinformatics, all of which are a barrier to many potential users. Here we present TITAN - an open-source, single environment, end-to-end pipeline that can be utilized for image visualization, segmentation, analysis and export of IMC data. TITAN is implemented as an extension within the publicly available 3D Slicer software. We demonstrate the utility, application, reliability and comparability of TITAN using publicly available IMC data from recently-published breast cancer and COVID-19 lung injury studies. Compared with current IMC analysis methods, TITAN provides a user-friendly, efficient single environment to accurately visualize, segment, and analyze IMC data for all users.

Effects of genotype and culture conditions on microspore embryogenesis in radish (Raphanus sativus L.).

Radish (Raphanus sativus L.), an important annual or biennial root vegetable crop, is widely cultivated in the world for its high nutritive value. Isolated microspore culture (IMC) is one of the most effective methods for rapid development of homozygous lines. Due to imperfection of the IMC technology system, it is particularly important to establish an efficient IMC system in radish. In this study, the effects of different factors on radish microspore embryogenesis were investigated with 23 genotypes. Buds with the largest population of late-uninucleate-stage microspores were most suitable for embryogenesis, with a ratio of petal length to anther length (P/A) in buds of about 3/4 ~ 1. Cold pretreatment was found to be genotype specific, and the highest microspore-derived embryoid (MDE) yield occurred for treatment of the heat shock of 48 h. In addition, the supplement of 0.75 g/L activated charcoal (AC) could increase the yield of embryoids. It was found that genotypes, bud size, as well as temperature treatments had significant effects on microspore embryogenesis. Furthermore, somatic embryogenesis-related kinase (SERK) genes were profiled by reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis, which indicated that they are involved in the process of MDE formation and plantlet regeneration. The ploidy of microspore-derived plants was identified by chromosome counting and flow cytometry, and the microspore-derived plants were further proved as homozygous plants through expressed sequence tags-simple sequence repeats (EST-SSR) and genetic-SSR markers. The results would facilitate generating the large-scale double haploid (DH) from various genotypes, and promoting further highly efficient genetic improvement in radish. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-022-01312-w.

Effects of probiotic Lactiplantibacillus plantarum IMC 510 supplementation on metabolic factors in otherwise healthy overweight and obese individuals.

AIMS: Probiotic supplementation approach offers the possibility to shape the gut microbiota (GM), enabling the development of innovative formulations able to improve intestinal well-being and consequently the related body weight modulation and energy metabolism. In the present clinical study, a new potential probiotic supplement based on Lactiplantibacillus plantarum IMC 510 was studied for weight management. METHODS AND RESULTS: Quantitative characterization by qPCR of representative bacterial groups of GM was used to determine the microbiota modulation at different supplementation periods. Furthermore, measurement of the endpoints linked to weight control (body mass index, body weight, waist circumference) was assessed. Specific questionnaires to evaluate the impact on psychological and physiological point of view were performed. Results showed that after 90 days, Lact. plantarum IMC 510 supplementation brought an improvement in endpoints linked to weight control and healthy status, although no significant changes in the microbiota composition were reported for analysed bacterial groups, except for Lactobacillus spp. and Bifidobacterium spp. CONCLUSIONS: We concluded that Lact. plantarum IMC 510 supplementation could be an interesting tool for weight management. More studies are needed to understand the impact on GM, for example, evaluating the production of short-chain fatty acids, since their important role in dietary metabolism. Further research is necessary to better elucidate the relationship between GM and overweight and the mechanism of action by which Lact. plantarum IMC 510 modifies body weight. SIGNIFICANCE AND IMPACT OF THE STUDY: However, these promising outcomes represent a clear advantage of probiotic supplementation and identify a new potential probiotic as a novel and safe therapeutic approach in the obesity prevention and management.

Engineering Tolerance toward Allogeneic CAR-T Cells by Regulation of MHC Surface Expression with Human Herpes Virus-8 Proteins.

Allogeneic, off-the-shelf (OTS) chimeric antigen receptor (CAR) cell therapies have the potential to reduce manufacturing costs and variability while providing broader accessibility to cancer patients and those with other diseases. However, host-versus-graft reactivity can limit the durability and efficacy of OTS cell therapies requiring new strategies to evade adaptive and innate-immune responses. Human herpes virus-8 (HHV8) maintains infection, in part, by evading host T and natural killer (NK) cell attack. The viral K3 gene encodes a membrane-tethered E3 ubiquitin ligase that discretely targets major histocompatibility complex (MHC) class I components, whereas K5 encodes a similar E3 ligase with broader specificity, including MHC-II and the MHC-like MHC class I polypeptide-related sequence A (MIC-A)- and sequence B (MIC-B)-activating ligands of NK cells. We created γ-retroviruses encoding K3 and/or K5 transgenes that efficiently transduce primary human T cells. Expression of K3 or K5 resulted in dramatic downregulation of MHC-IA (human leukocyte antigen [HLA]-A, -B, and -C) and MHC class II (HLA-DR) cell-surface expression. K3 expression was sufficient for T cells to resist exogenously loaded peptide-MHC-specific cytotoxicity, as well as recognition in one-way allogeneic mixed lymphocyte reactions. Further, in immunodeficient mice engrafted with allogeneic T cells, K3-transduced T cells selectively expanded in vivo. Ectopic K5 expression in MHC class I-, MIC-A+/B+ K562 cells also reduced targeting by primary NK cells. Coexpression of K3 in prostate stem cell antigen (PSCA)-directed, inducible MyD88/CD40 (iMC)-enhanced CAR-T cells did not impact cytotoxicity, T cell growth, or cytokine production against HPAC pancreatic tumor target cells, whereas K5-expressing cells showed a modest reduction in interleukin (IL)-2 production without effect on cytotoxicity. Together, these results support application of these E3 ligases to advance development of OTS CAR-T cell products.

A phase I study of IMC-001, a PD-L1 blocker, in patients with metastatic or locally advanced solid tumors.

Introduction IMC-001 is a fully human IgG1 monoclonal antibody that binds to human PD-L1 (programmed death-ligand 1). This study evaluated the safety, pharmacokinetics, and pharmacodynamics of IMC-001 in patients with advanced solid tumors. Materials and Methods This open-labeled phase I study used a standard 3 + 3 dose-escalation design, with doses ranging from 2 to 20 mg/kg. IMC-001 was administered intravenously every 2 weeks until disease progression or unacceptable toxicity. The dose-limiting toxicity window was defined as 21 days from the first dose. Results Fifteen subjects were included in 5 dose-escalation cohorts. No dose-limiting toxicity was observed, and the maximum tolerated dose was not reached. The most common adverse events (AEs) were general weakness, decreased appetite, fever, and cough. No grade 4 or 5 treatment emergent AEs were reported during the study. One subject in the 2 mg/kg cohort showed grade 2 immune-induced thyroiditis and diabetes mellitus suspected to be related to IMC-001. Over the dose range of 2-20 mg/kg IMC-001, the AUC0-14d, AUC0-∞, and Cmax generally increased in a dose-proportional manner for each step of dose escalation. Of the 15 enrolled patients, 1 subject with rectal cancer showed a partial response, and the disease control rate was 33.3%. Conclusions IMC-001 demonstrated a favorable safety profile up to 20 mg/kg administered intravenously every 2 weeks and showed preliminary efficacy in patients with advanced solid tumors. Based on pharmacokinetic and pharmacodynamic data, 20 mg/kg was selected as the recommended phase II dose. Clinical trial identification NCT03644056 (date of registration: August 23, 2018).

Association of blood pressure and BMI to corrected QT interval in young adults.

Hypertension is often associated with obesity. Uncontrolled hypertension can lead to uncorrected cardiac dysautonomia that makes cardiac repolarization abnormally prolonged. Modern dietary habits, stress, and bad lifestyle habits make young adults vulnerable to hazards of health. We planned to study the association of body mass index (BMI) and blood pressure (BP) with corrected QT (QTc) intervals in young adults. After obtaining the written informed consent, 171 subjects were randomly selected in the age group of 18-35 years. A general history and physical examination were done before recording the BMI and BP. A 12-lead electrocardiogram was recorded and QTc calculated using Bazett's formula. The values obtained were compared and statistical analysis done. Of the 171 subjects 14.03% were hypertensive. The QTc interval was found to be prolonged in females and it is prolonged with age, BMI, and BP. QTc was significantly increased among the prehypertensive and hypertensive group and with the overweight and (or) obese BMI group compared with the normotensives and normal BMI group suggesting an altered autonomic homeostasis. This warrants lifestyle modification at a younger age to reduce the cardiovascular risk.

Imaging mass cytometry for high-dimensional tissue profiling in the eye.

BACKGROUND: Imaging mass cytometry (IMC) combines the principles of flow cytometry and mass spectrometry (MS) with laser scanning spatial resolution and offers unique advantages for the analysis of tissue samples in unprecedented detail. In contrast to conventional immunohistochemistry, which is limited in its application by the number of possible fluorochrome combinations, IMC uses isoptope-coupled antibodies that allow multiplex analysis of up to 40 markers in the same tissue section simultaneously. METHODS: In this report we use IMC to analyze formalin-fixed, paraffin-embedded conjunctival tissue. We performed a 18-biomarkers IMC analysis of conjunctival tissue to determine and summarize the possibilities, relevance and limitations of IMC for deciphering the biology and pathology of ocular diseases. RESULTS: Without modifying the manufacturer's protocol, we observed positive and plausible staining for 12 of 18 biomarkers. Subsequent bioinformatical single-cell analysis and phenograph clustering identified 24 different cellular clusters with distinct expression profiles with respect to the markers used. CONCLUSIONS: IMC enables highly multiplexed imaging of ocular samples at subcellular resolution. IMC is an innovative and feasible method, providing new insights into ocular disease pathogenesis that will be valuable for basic research, drug discovery and clinical diagnostics.

Inertial Motion Capture-Based Whole-Body Inverse Dynamics.

Inertial Motion Capture (IMC) systems enable in situ studies of human motion free of the severe constraints imposed by Optical Motion Capture systems. Inverse dynamics can use those motions to estimate forces and moments developing within muscles and joints. We developed an inverse dynamic whole-body model that eliminates the usage of force plates (FPs) and uses motion patterns captured by an IMC system to predict the net forces and moments in 14 major joints. We validated the model by comparing its estimates of Ground Reaction Forces (GRFs) to the ground truth obtained from FPs and comparing predictions of the static model's net joint moments to those predicted by 3D Static Strength Prediction Program (3DSSPP). The relative root-mean-square error (rRMSE) in the predicted GRF was 6% and the intraclass correlation of the peak values was 0.95, where both values were averaged over the subject population. The rRMSE of the differences between our model's and 3DSSPP predictions of net L5/S1 and right and left shoulder joints moments were 9.5%, 3.3%, and 5.2%, respectively. We also compared the static and dynamic versions of the model and found that failing to account for body motions can underestimate net joint moments by 90% to 560% of the static estimates.

Conceptual Design and Image Motion Compensation Rate Analysis of Two-Axis Fast Steering Mirror for Dynamic Scan and Stare Imaging System.

In order to enable the aerial photoelectric equipment to realize wide-area reconnaissance and target surveillance at the same time, a dual-band dynamic scan and stare imaging system is proposed in this paper. The imaging system performs scanning and pointing through a two-axis gimbal, compensating the image motion caused by the aircraft and gimbal angular velocity and the aircraft liner velocity using two two-axis fast steering mirrors (FSMs). The composition and working principle of the dynamic scan and stare imaging system, the detailed scheme of the two-axis FSM and the image motion compensation (IMC) algorithm are introduced. Both the structure and the mirror of the FSM adopt aluminum alloys, and the flexible support structure is designed based on four cross-axis flexural hinges. The Root-Mean-Square (RMS) error of the mirror reaches 15.8 nm and the total weight of the FSM assembly is 510 g. The IMC rate equations of the two-axis FSM are established based on the coordinate transformation method. The effectiveness of the FSM and IMC algorithm is verified by the dynamic imaging test in the laboratory and flight test.

Supplementation with Lactiplantibacillus plantarum IMC 510 Modifies Microbiota Composition and Prevents Body Weight Gain Induced by Cafeteria Diet in Rats.

Changes in functionality and composition of gut microbiota (GM) have been associated and may contribute to the development and maintenance of obesity and related diseases. The aim of our study was to investigate for the first time the impact of Lactiplantibacillus (L.) plantarum IMC 510 in a rat model of diet-induced obesity, specifically in the cafeteria (CAF) diet. This diet provides a strong motivation to voluntary overeat, due to the palatability and variety of selected energy-dense foods. The oral administration for 84 days of this probiotic strain, added to the CAF diet, decreased food intake and body weight gain. Accordingly, it ameliorated body mass index, liver and white adipose tissue weight, hepatic lipid accumulation, adipocyte size, serum parameters, including glycemia and low-density lipoprotein levels, in CAF fed rats, potentially through leptin control. In this scenario, L. plantarum IMC 510 showed also beneficial effects on GM, limiting the microbial imbalance established by long exposure to CAF diet and preserving the proportion of different bacterial taxa. Further research is necessary to better elucidate the relationship between GM and overweight and then the mechanism of action by which L. plantarum IMC 510 modifies weight. However, these promising results prompt a clear advantage of probiotic supplementation and identify a new potential probiotic as a novel and safe therapeutic approach in obesity prevention and management.

U-Model-Based Two-Degree-of-Freedom Internal Model Control of Nonlinear Dynamic Systems.

This paper proposes a U-Model-Based Two-Degree-of-Freedom Internal Model Control (UTDF-IMC) structure with strength in nonlinear dynamic inversion, and separation of tracking design and robustness design. This approach can effectively accommodate modeling error and disturbance while removing those widely used linearization techniques for nonlinear plants/processes. To assure the expansion and applications, it analyses the key properties associated with the UTDF-IMC. For initial benchmark testing, computational experiments are conducted using MATLAB/Simulink for two mismatched linear and nonlinear plants. Further tests consider an industrial system, in which the IMC of a Permanent Magnet Synchronous Motor (PMSM) is simulated to demonstrate the effectiveness of the design procedure for potential industrial applications.

Rare osteosarcoma cell subpopulation protein array and profiling using imaging mass cytometry and bioinformatics analysis.

BACKGROUND: Single rare cell characterization represents a new scientific front in personalized therapy. Imaging mass cytometry (IMC) may be able to address all these questions by combining the power of MS-CyTOF and microscopy. METHODS: We have investigated this IMC method using < 100 to up to 1000 cells from human sarcoma tumor cell lines by incorporating bioinformatics-based t-Distributed Stochastic Neighbor Embedding (t-SNE) analysis of highly multiplexed IMC imaging data. We tested this process on osteosarcoma cell lines TC71, OHS as well as osteosarcoma patient-derived xenograft (PDX) cell lines M31, M36, and M60. We also validated our analysis using sarcoma patient-derived CTCs. RESULTS: We successfully identified heterogeneity within individual tumor cell lines, the same PDX cells, and the CTCs from the same patient by detecting multiple protein targets and protein localization. Overall, these data reveal that our t-SNE-based approach can not only identify rare cells within the same cell line or cell population, but also discriminate amongst varied groups to detect similarities and differences. CONCLUSIONS: This method helps us make greater inroads towards generating patient-specific CTC fingerprinting that could provide an accurate tumor status from a minimally-invasive liquid biopsy.