Visual Quantification Research Articles

Sustainable Utilization of Machine-Vision-Technique-Based Algorithm in Objective Evaluation of Confocal Microscope Images

Confocal microscope imaging has become popular in biotechnology labs. Confocal imaging technology utilizes fluorescence optics, where laser light is focused onto a specific spot at a defined depth in the sample. A considerable number of images are produced regularly during the process of research. These images require methods of unbiased quantification to have meaningful analyses. Increasing efforts to tie reimbursement to outcomes will likely increase the need for objective data in analyzing confocal microscope images in the coming years. Utilizing visual quantification methods to quantify confocal images with naked human eyes is an essential but often underreported outcome measure due to the time required for manual counting and estimation. The current method (visual quantification methods) of image quantification is time-consuming and cumbersome, and manual measurement is imprecise because of the natural differences among human eyes’ abilities. Subsequently, objective outcome evaluation can obviate the drawbacks of the current methods and facilitate recording for documenting function and research purposes. To achieve a fast and valuable objective estimation of fluorescence in each image, an algorithm was designed based on machine vision techniques to extract the targeted objects in images that resulted from confocal images and then estimate the covered area to produce a percentage value similar to the outcome of the current method and is predicted to contribute to sustainable biotechnology image analyses by reducing time and labor consumption. The results show strong evidence that t-designed objective algorithm evaluations can replace the current method of manual and visual quantification methods to the extent that the Intraclass Correlation Coefficient (ICC) is 0.9.

Ratiometric dual-emission carbon dots coupled with smartphone for visual quantification of Co2+ and EDTA and biological sensing

Ratiometric fluorescence analysis provides the feasibility for visual quantitative sensing by presenting continuous color changes. Herein, dual-emission carbon dots (D-CDs) have been developed for ratiometric recognition of Co2+ and EDTA, along with successive fluorescence variation of yellow, purplish-pink, purple, light blue, and dark blue. D-CDs have been synthesized by microwave-assisted heating of tetracycline and manifest dual emissions at 562 nm and 610 nm under 450 nm excitation. Intriguingly, the emissions at 562 nm and 610 nm are quenched while a ratio rising emission emerges at 482 nm upon introducing Co2+, which displays ratiometric fluorescence emission characteristics (I482/I562) in the range of 0.25–400 µM with significant fluorescence varying from yellow, purplish-pink, purple, light blue, to dark blue. Subsequent addition of EDTA could ratiometric recover the fluorescence of D-CDs in the range of 50–130 µM with the fluorescence returning from dark blue, light blue, purple, purplish-pink, to yellow. More crucially, integrating continuous color changes and color recognizer APP in the smartphone, visual quantification of Co2+ and EDTA can be realized based on the ratio of blue value and red value (B/R) with linear ranges of 0–400 µM and 0–300 µM, respectively. Furthermore, proposed D-CDs could be extended as a visual biosensing platform for tracking Co2+ and EDTA in living cells, which have potentialapplication prospect in biosystem.

Ion-induced enhanced fluorescence colorimetric hydrogel sensor for visual quantization of doxycycline

The potential hazard to human health caused by antibiotic residues demands a rapid and sensitive detection method. However, the current rapid methods, such as paper-based sensors with reduced detection sensitivity caused by probe solidification due to printing, affect detection accuracy. Here, a hydrogel sensor has been prepared by an ion-induced enhanced optical probe using lutetium ions (Lu3+) coordinated with CdTe quantum dots (QDs). The hydrogel integrates the advantages of the liquid-phase in- situ detection environment and higher accuracy. With the addition of doxycycline (Dox), the β-diketone structure of Dox forms complexes with Lu3+, leading to a change in the Dox molecular structure, and absorbs electrons from QDs that provide a significant color change from red to green. The probe reacts within 2 s with a limit of detection (LOD) of 36.6 nM. Integrated with the advantages of the hydrogel and color analyzer app, the LOD achieved 53.7 nM, which is close to the detection sensitivity in water. On this basis, a smartphone sensing platform has been constructed for the rapid, visual and quantitative analysis of Dox. The hydrogel-based sensing method for rapid visual quantification detection of Dox provides a new strategy for food safety detection.

Identification of Redox Shuttle Generated in LFP/Graphite and NMC811/Graphite Cells

Unwanted self-discharge of LFP/AG and NMC811/AG cells can be caused by in situ generation of a redox shuttle molecule after formation at elevated temperature with common alkyl carbonate electrolyte. This study investigates the redox shuttle generation for several electrolyte additives, e.g., vinylene carbonate and lithium difluorophosphate, by measuring the additive reduction onset potential, first cycle inefficiency and gas evolution during formation at temperatures between 25 and 70 °C. After formation, electrolyte is extracted from pouch cells for visual inspection and quantification of redox shuttle activity in coin cells by cyclic voltammetry. The redox shuttle molecule is identified by GC-MS and NMR as dimethyl terephthalate. It is generated in the absence of an effective SEI-forming additive, according to a proposed formation mechanism that requires residual water in the electrolyte, catalytic quantities of lithium methoxide generated at the negative electrode and, surprisingly, polyethylene terephthalate tape within the cell.

Point of care testing (POCT) of cholesterol in blood serum via a moving reaction boundary electrophoresis titration chip.

Cholesterol (CHO) in human blood is one of the most frequently and crucially quantified substances in diagnostic laboratories. However, visual and portable point of care testing (POCT) methods have been rarely developed for the bioassay of CHO in blood samples. Here, we developed an electrophoresis titration (ET) model, a chip device of ∼60 grams, and a quantification method for the POCT of CHO in blood serum based on a moving reaction boundary (MRB). In this model, the selective enzymatic reaction is integrated with an ET chip for visual and portable quantification. At first, CHO reacted with cholesterol oxidase (CHOx) in the anode well, producing H2O2 and cholest-4-en-3-one in the solution. H2O2 further oxidized the colorless and chargeless leucocrystal violet (LCV) dye into violet colored positively charged crystal violet (CV+) and, under the influence of the electric field, the CV+ migrates in the ET channels and is titrated by the alkali of sodium hydroxide immobilized in the ET channels. The length covered by the MRB was measured as a function of the CHO content. The relevant experiments validated the feasibility of the model and method. Furthermore, the experiments revealed the high selectivity, portability, and visuality of the ET-MRB model, device, and method. Finally, the experiments showed a fair sensitivity of LOD of 5 μM, good linearity of 10-1000 μM (r2 = 0.9919), fair stability (intra-day RSD of less than 5.09% and an inter-day RSD of less than 6.36%), and high recovery (99.4-105%). All the data and results indicate the potential of the ET-MRB model, chip device, and method for POCT of CHO in human blood samples.

Visual quantification of prostaglandin E2 discharge from a single cell.

Calcium transients drive cells to discharge prostaglandin E2 (PGE2). We visualized PGE2-induced protein kinase A (PKA) activation and quantitated PGE2 secreted from a single cell by combining fluorescence microscopy and a simulation model. For this purpose, we first prepared PGE2-producer cells that express either an optogenetic or a chemogenetic calcium channel stimulator: OptoSTIM1 or Gq-DREADD, respectively. Second, we prepared reporter cells expressing the Gs-coupled PGE2 reporter EP2 and the PKA biosensor Booster-PKA, which is based on the principle of Förster resonance energy transfer (FRET). Upon the stimulation-induced triggering of calcium transients, a single producer cell discharges PGE2 to stimulate PKA in the surrounding reporter cells. Due to the flow of the medium, the PKA-activated area exhibited a comet-like smear when HeLa cells were used. In contrast, radial PKA activation was observed when confluent MDCK cells were used, indicating that PGE2 diffusion was restricted to the basolateral space. By fitting the radius of the PKA-activated area to a simulation model based on simple diffusion, we estimated that a single HeLa cell secretes 0.25 fmol PGE2 upon a single calcium transient to activate PKA in more than 1000 neighboring cells. This model also predicts that the PGE2 discharge rate is comparable to the diffusion rate. Thus, our method quantitatively envisions that a single calcium transient affects more than 1000 neighboring cells via PGE2.Key words: prostaglandin E2, imaging, intercellular communication, biosensor, quantification.

ВЫБОР МЕТОДА РЕКОНСТРУКЦИИ ПОЗИТРОННО-ЭМИССИОННЫХ ЭКГ-СИНХРОНИЗОВАННЫХ ИССЛЕДОВАНИЙ И ЕГО ВЛИЯНИЕ НА РАССЧИТАННЫЕ ОБЪЁМНЫЕ ПОКАЗАТЕЛИ ФУНКЦИИ МИОКАРДА ЛЕВОГО ЖЕЛУДОЧКА И ВИЗУАЛЬНОЕ КАЧЕСТВО ИЗОБРАЖЕНИЙ

Ischemic heart disease is one of the leading causes of mortality in the Russian Federation. Timely diagnosis of this condition allows you to determine the optimal treatment tactics. Positron emission tomography with 18-fluorodeoxyglucose is a method for verifying viable cardiac tissue in patients with myocardial infarction. The methods used for the reconstruction of positron emission tomograms can affect the quantitative indicators of the contractile function of the myocardium. Materials and methods. The study retrospectively included 12 patients who underwent PET/CT with 18-fluorodeoxyglucose, the results of which were reconstructed using filtered back projection methods and iterative reconstruction, with ECG synchronization and division of ECG R-R intervals into 8, 10 and 16 frames. The obtained images were evaluated visually by two radiologists, and the quantitative parameters characterizing the contractile function of the left ventricular myocardium were compared in pairs. Results. There were no significant differences in the assessment of quantitative indicators such as end-systolic and diastolic volumes, stroke volume and ejection fraction. However, the iterative reconstruction results show less data scatter than the filtered back projection reconstruction method. In visual analysis, the best quality of reconstructions by iterative methods is noted. Conclusions. Thus, the recommended method for both visual analysis and quantification is iterative reconstruction, regardless of the number of timing windows used. A spread in the number of used reconstructed fragments of the cardiac cycle in the range of 8-16, commonly used in clinical practice, can be recommended.

Quantification analysis of microplastics released from disposable polystyrene tableware with fluorescent polymer staining

Ingesting microplastics (MPs) from plastic tableware is an important source of health risk to human bodies. However, the comprehensive information of MPs released from disposable tableware has not been explored. Herein, a new visual quantification method for polystyrene MPs is proposed with carbon nitride fluorescent polymers staining, which can overcome the disadvantages of high signal background and photobleaching derived from organic dyes staining. Combining with fluorescence microscope and ImageJ software, the quantity, shape, and size distribution of MPs carried by the brand-new disposable polystyrene tableware (DPT) samples before usage and released from the clean DPT samples in different simulated usage scenes were studied. The brand-new DPT samples were found to carry a large number of MPs particles and the clean DPT samples could release MPs during usage. Fiber and fragment are the main morphology of the detected MPs and fiber accounts for 45–52 %. The particles with size <50 μm are the majority of the detected MPs and the distribution fraction of MPs particles is gradually decreased with the raising of particle size within 50 μm. The released MPs particles are increased with the raising of contact time and temperature, and greatly boosted for the DPT samples with cracks. The DPT samples are more like to release MPs in weak acidic condition (pH 4.0) than in weak alkaline (pH 8.3) and neutral (pH 7.0) conditions. The obtained results help to assess the food safety of tack-out food and the health risk of MPs exposure to human.

Visual, spectral, and microchemical quantification of crystalline anomalies in otoliths of wild and cultured delta smelt

Developmental abnormalities in otoliths can impact growth and survival in teleost fishes. Here, we quantified the frequency and severity of developmental anomalies in otoliths of delta smelt (Hypomesus transpacificus), a critically endangered estuarine fish that is endemic to the San Francisco Estuary. Left–right asymmetry and anomalous crystalline polymorphs (i.e., vaterite) were quantified and compared between wild and cultured populations using digital image analysis. Visual estimates of vaterite were validated using X-ray diffraction, Raman spectroscopy, laser ablation ICPMS, and electron probe microanalysis. Results indicated that cultured delta smelt were 80 times more likely to contain a vateritic otolith and 18 times more likely to contain relatively large (≥ 15%) amounts of vaterite. Similarly, cultured fish exhibited 30% greater asymmetry than wild fish. These results indicate that cultured delta smelt exhibit a significantly higher frequency of vestibular abnormalities which are known to reduce fitness and survival. Such hatchery effects on otolith development could have important implications for captive culture practices and the supplementation of wild fish populations with cultured individuals.

A Fully Integrated, Ready-to-Use Distance-Based Chemosensor for Visual Quantification of Multiple Heavy Metal Ions.

Point-of-care devices offering quantitative results with simple steps would allow great useability for untrained end-users. Here, we report a ready-to-use chemosensor integrating automatic sample metering, on-chip reaction, gravitational-magnetic separation, and a distance-based readout for visual quantification of multiple heavy metal ions. Deoxyribozymes (DNAzymes), probe-modified magnetic microparticles (MMPs), and polystyrene microparticles (PMPs) are preloaded into a microfluidic chip and freeze-dried. After the water sample is collected with automatic volume metering, the particles are resuspended, and the MMPs and PMPs hybridize with DNAzyme at its two termini, forming the "MMPs-DNAzyme-PMPs" structure. When target metal ions are present, the DNAzymes are cleaved, yielding an increased number of free PMPs. All on-chip reactions are controlled by stopping the liquid flow at capillary valves and bursting it with hand-controlled tilting. Using the chip with a gravitational-magnetic separator, the free PMPs are separated from "MMPs-DNAzyme-PMPs" and accumulate into the trapping channel with a nozzle, forming a visual bar with growing distances proportional to the concentration of target metal ions. The achieved limit of detection (LOD) values for Cu2+ (103.1 nM), Pb2+ (69.5 nM), and Ag+ (793.6 nM) are below the maximum contamination levels. High selectivity of 100-fold, 200-fold, and 20-fold against interference is obtained. Moreover, by integrating three identical channels in parallel, simultaneous detection of the above-mentioned heavy metal ions in fresh and tap water samples is also achieved with high accuracy. Together, this fully integrated and easily operated platform embodies excellent potential for rapid, on-site sensing by unskilled users.

Microplastic contamination in commercial salt: An issue for their sampling and quantification

Sea salt can be considered as a vector of microplastics in the human body. In this work, the sea salts collected from three Italian salterns has been solubilized in MilliQ water and filtered to extract microplastics. The visual quantification of microplastics with a stereomicroscope was carried out on the bases of their size, followed by a classification taking into account their physical characteristics. ATR-FTIR and Raman spectroscopy were used to identify the polymeric type of microplastics. Their significant presence has been revealed: 1653 ± 29 microplastics/kg of sea salt. In total, 80.6 % of microplastics have a fiber shape, 18.9 % a fragmented shape and 2.7 % are sphere. The size of microplastics has been analysed, indicating that the most frequent is between 0 and 500 µm. Polypropylene, polyamide and polyethylene were identified as the most frequent types of polymers. This research could be of global relevance given the significant export of Italian salt to foreign countries.

Enzyme-nanozyme cascade colorimetric sensor platform: a sensitive method for detecting human serum creatinine.

Creatinine, as a significant biomarker for kidney, thyroid, and muscle dysfunction-related diseases, detection is of great important meaning. In this paper, an enzyme-nanozyme cascade sensing platform was developed for visual creatinine detection. Perovskite oxide BiFeO3 synthesized by a sol-gel method was applied as a nanozyme, showing excellent peroxidase-like activity. During detection, creatinine was oxidized in turn by three natural enzymes (creatinase, creatininase, and sarcosine oxidase) to produce H2O2, and H2O2 was then catalyzed by the BiFeO3 nanozyme, resulting in the change of achromatous 3,3',5,5'-tetramethylbenzidine (TMB) into blue oxidized TMB (oxTMB). Based on this principle, visual quantification of creatinine was realized. Due to the high stability and catalytic efficiency of nanozyme, the cascade sensing platform can be used to detect creatinine in a broad range of 0.5-150μM with a detection limit of 0.09μM. Meanwhile, thanks to the specificity of the natural enzymes, the platform exhibited admirable selectivity for creatinine determination despite the existence of a variety of interfering substances, which were successfully adopted to measure the level of creatinine in human serums. The cascade sensing platform is expected to serve the determination of a large number of biomarkers by simply alternating the natural enzymes.

Abstract 627: Custom Vectra® Polaris™ fluorescent multiplex IHC panel identifies mature tertiary lymphoid structures in colorectal cancer

Abstract Introduction: Tertiary lymphoid structures (TLS) are promising prognostic indicators of positive outcomes for patients with solid tumors including colorectal cancer (CRC) [1]. Large-scale retrospective analysis shows patients with mature TLS in particular respond to PD-1/PD-L1 antibody treatment with improved objective response, progression-free and overall survival [2]. Since not all patients respond to PD-1/PD-L1 antibody treatment, identifying patients with mature TLS is clinically relevant as it enables selection of patients likely to respond to immune checkpoint blockade. Mature TLS are composed of T cells (CD3+), follicular B cells (CD20+), germinal center B cells (CD23+), and follicular dendritic cells (CD21+) [3]. The ability to identify and evaluate TLS is limited by detection methods which traditionally employ hematoxylin and eosin (H&amp;E) staining for visual quantification of immune aggregates, potentially significantly underestimating their quantity [4]. Multiplexed fluorescent immunohistochemistry (IHC) detection assays have the capability to precisely quantify mature TLS within the TME. Colorectal cancer specimens expressing a dynamic range of TLS are stained with a novel custom Vectra® Polaris™ multiplex immunofluorescence panel detecting CD20, CD21, CD23, CD3 and pan-cytokeratin. Identification of mature TLS with CD3+CD20+CD21+ and CD3+CD20+CD23+ expression in the TME is reported. Results: Vectra® Polaris™ TLS multiplex immunofluorescence panel successfully identified mature TLS in the TME of colorectal cancer patient samples. TLS were quantified via custom analytics algorithms generated with Indica HALO software. Conclusion: Characteristics of TME immunity in colorectal cancer differentially impact an individual patient’s odds of survival [5]. The novel Vectra® Polaris™ multiplex assay in this study shows a detailed picture of mature TLS in patient cancer samples. Future applications of this panel include investigations of TLS associated with successful anti-tumor immune development and therapeutic response to treatment. [1] Oncoimmunology. 2020; 9(1): 1724763 [2] Nat Cancer. 2, 794-802 (2021) [3] Oncoimmunology. 2021; 10(1): 1900508. [4] Mod Pathol 2017 Sep;30(9):1204-1212. doi: 10.1038/modpathol.2017.43. [5] Cell. 2020 Sep 3;182(5):1341-1359.e19. doi: 10.1016/j.cell.2020.07.005 Citation Format: Sara G. Pollan, Arezoo Hanifi, James Hargrove, Erinn A. Parnell, Jessica Lin, Josette William Ragheb, Qingyan Au. Custom Vectra® Polaris™ fluorescent multiplex IHC panel identifies mature tertiary lymphoid structures in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 627.

Electrochemical/visual microfluidic detection with a covalent organic framework supported platinum nanozyme-based device for early diagnosis of pheochromocytoma

The development of smart, portable, and sensitive devices for the monitoring of circulating tumor cells (CTCs) is essential to diagnose several diseases, including pheochromocytoma (PCC). Therefore, in this study, a dual-mode (electrochemical/visual) microfluidic device was designed for the rapid and sensitive detection of PCC-CTCs using a microfluidic chip for automatic cell sampling and detection and a smartphone-based three-dimensional-printed accessory for signal output analysis. The device was employed to capture and identify PCC-CTCs via specific immunogenic binding to the norepinephrine transporter and somatostatin receptor, which are overexpressed on the surface of PCC cells. Specifically, targeted-modified magnetic particles were used to capture and separate PCC-CTCs from peripheral blood; then, similarly modified covalent organic framework based nanozymes (COF@Pt) were used as peroxidase mimics to amplify the electrochemical response from H2O2 reduction and catalyze the oxidation of 3,3′,5,5′-tetramethylbenzidine by hydroxyl radicals in the presence of the PCC cells to enable visual quantification. Using the prepared microfluidic device, a low detection limit of 1 cell mL−1 at a signal-to-noise ratio of 3 and a wide linear range of 2 to 105 cells mL−1 were achieved. Overall, this work demonstrates a portable, sensitive, and visual platform for PCC diagnostics that meets the requirement for quick and precise point-of-care diagnostics.

COVIR: A virtual rendering of a novel NN architecture O-Net for COVID-19 Ct-scan automatic lung lesions segmentation

With the Coronavirus disease 2019 (COVID-19) spread, causing a world pandemic, and recently, the virus new variants continue to appear, making the situation more challenging and threatening, the visual assessment and quantification by expert radiologists have become costly and error-prone. Hence, there is a need to propose a model to predict the COVID-19 cases at the earliest possible to control the disease spread. In order to assist the medical professionals and reduce workload and the time the COVID-19 diagnosis cycle takes, this paper proposes a novel neural network architecture termed as O-Net to automatically segment chest Computerised Tomography Ct-scans infected by COVID-19 with optimised computing power and memory occupation. The O-Net consists of two convolutional autoencoders with an upsampling channel and a downsampling channel. Experimental tests show our proposal’s effectiveness and potential, with a dice score of 0.86, pixel accuracy, precision, specificity of 0.99, 0.99, 0.98, respectively. Performance on the external dataset illustrates generalisation and scalability capabilities of the O-Net model to Ct-scan obtained from different scanners with different sizes. The second objective of this work is to introduce our virtual reality platform, COVIR, that visualises and manipulates 3D reconstructed lungs and segmented infected lesions caused by COVID-19. COVIR platform acts as a reading and visualisation support for medical practitioners to diagnose COVID-19 lung infection. The COVIR platform could be used for medical education professional practice and training. It was tested by Thirteen participants (medical staff, researchers, and collaborators), they conclude that the 3D VR visualisation of segmented Ct-Scan provides an aid diagnosis tool for better interpretation.

Colorimetric aptasensor for on-site detection of acetamiprid with hybridization chain reaction-assisted amplification and smartphone readout strategy

An instruments-free colorimetric assay is established for acetamiprid detection with hybridization chain reaction (HCR)-based amplification strategy. Hairpin probes help gold nanoparticles (AuNPs) to resist salt-induced aggregation through sticky ends. In the presence of acetamiprid, aptamer hairpin probe HP opens due to the specifical recognition, then triggers an explosive hybridization between hairpin probes HA-1/2 and numerous hairpin probes are consumed. With the depletion of hairpin probes, the protection weakens and AuNPs suffer from salt-induced aggregation, which results in a color change from pinkish purplish to blue-purple. Under optimal conditions, the assay exhibits high sensitivity for acetamiprid analysis with a detection limit at 1.74 μM. Meanwhile, the aptasensor displays excellent selectivity that only acetamiprid can trigger the HCR and achieve enhanced detection. Worthwhile, based on the color variation, a smartphone readout-based strategy is proposed for visual quantification of acetamiprid, and good practical performance is obtained in real agricultural samples. The above visual method is portable and economic that get rid of advanced instruments, and shows a promising application prospect for on-site detection of acetamiprid.

Abstract P1-02-13: Tumour infiltrating lymphocytes (TILs) and immune composition in breast cancer patients from Kenya: Spatial distributions and associations with risk factors and tumour characteristics

Abstract Background: The immune landscape of breast cancer in patients from Sub Saharan Africa is understudied and findings are mainly extrapolated from studies in Caucasians and African Americans. Given the prognostic value of Tumour Infiltrating Lymphocytes (TILs) in breast cancer subtypes, the paucity of specific TIL marker data among BC patients from Africa, and the limited data on TIL spatial heterogeneity, our aim was to describe the distribution of TILs within the intratumoral stroma and the leading/invasive edge stroma, and evaluate their association across BC subtypes with established risk factors in Kenyan women with BC. Methods: Visual quantification of overall TILs within the intratumoral stroma (sTILs) and the stroma at the leading edge/invasive edge of the tumour (LE-TILs) were performed on Haematoxylin and Eosin stained whole slide sections of pathologically confirmed breast cancer based on the guidelines of the International TIL working group. Tissue Microarrays (TMAs) of tumour were also were constructed and stained with immunohistochemistry for CD3, CD4, CD8, CD68, CD20 and Fox P3. The TMA slides were scanned on the Aperio Digital pathology slide scanner, imported into the Aperio, assigned a template, segmented and annotated before analysis using a nuclear algorithm. Linear and logistic regression models were used to assess the associations between risk factors and tumor features with IHC markers and total TILs, after adjusting for other covariates. Results: A total of 226 invasive breast cancer cases were included in the final analysis for TILs and immune cell markers. The mean age at diagnosis of these breast cancer patients was 48.3 years. The distribution of the BC molecular subtypes defined by IHC markers was 56.2%, 16.8%, 9.3%, and 17.7 % (luminal A, luminal B, HER2-enriched, and TN breast cancers, respectively). On the whole we found that LE TIL proportions were higher than sTIL (41.3% versus 15% for TILs ≥30 and 19.3% and 4.4% for TILs ≥50 respectively). After adjusting for other tumor characteristics, we found higher TILs associated with high KI67 and high grade (for both sTIL and LE-TIL), TNBC (for sTIL only), and HER2 status, luminal B subtype, and smaller tumor size (&amp;lt;4 cm) (for LE-TIL only). Both sTILs and LE-TILs were predominantly composed of CD3, CD8 and CD68 with a much smaller contribution of CD4 and CD20. The percent positive cells for Fox3 were markedly lower compared to other IHC markers for both sTILs and LE-TILs. Conclusions: Our findings with regards to enrichment of TILs in more aggressive breast cancers are similar to what has been previously published. The distinct associations between intratumoral stroma and leading edge TIL measures for some factors highlight the importance of spatial TIL evaluations in future studies. Our findings add to the body of knowledge regarding TILs and TIL subtypes in breast cancer, which underscore the need to conduct studies specific to African populations. Citation Format: Shahin Sayed, Xiaohong R Yang, Dhirendra Govender, Breast Health Study group. Tumour infiltrating lymphocytes (TILs) and immune composition in breast cancer patients from Kenya: Spatial distributions and associations with risk factors and tumour characteristics [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P1-02-13.

COVID-19 CT Scan Lung Segmentation: How We Do It.

The National Health Systems have been severely stressed out by the COVID-19 pandemic because 14% of patients require hospitalization and oxygen support, and 5% require admission to an Intensive Care Unit (ICU). Relationship between COVID-19 prognosis and the extent of alterations on chest CT obtained by both visual and software-based quantification that expresses objective evaluations of the percentage of ventilated lung parenchyma compared to the affected one has been proven. While commercial applications for automatic medical image computing and visualization are expensive and limited in their spread, the open-source systems are characterized by not enough standardization and time-consuming troubles. We analyzed chest CT exams on 246 patients suspected of COVID-19 performed in the Emergency Department CT room. The lung parenchyma segmentation was obtained by a threshold-based method using the open-source 3D Slicer software and software tools called “Segment Editor” and “Segment Quantification.” For the three main characteristics analyzed on lungs affected by COVID-19 pneumonia, a specifical densitometry value range was defined: from − 950 to − 700 HU for well-aerated parenchyma; from − 700 to − 250 HU for interstitial lung disease; from − 250 to 250 HU for parenchymal consolidation. For the well-aerated parenchyma and the interstitial alterations, the procedure was semi-automatic with low time consumption, whereas consolidations’ analysis needed manual interventions by the operator. After the chest CT, 13% of the sample was admitted to intensive care, while 34% of them to the sub-intensive care. In patients moved to intensive care, the parenchyma analysis reported a higher crazy paving presentation. The quantitative analysis of the alterations affecting the lung parenchyma of patients with COVID-19 pneumonia can be performed by threshold method segmentation on 3D Slicer. The segmentation could have an important role in the quantification in different COVID-19 pneumonia presentations, allowing to help the clinician in the correct management of patients.

Evaluation of a 3D-printed pipette tip for seeding housefly eggs

Egg manipulation is crucial in insect breeding and research. Housefly eggs are small, delicate and susceptible to dehydration. Hence, researchers require rapid and gentle ways to manipulate specific quantities, with consistent quality. Therefore, a 3D-printed pipette tip was designed and tested. Three series of pipette tips, differing in volume (A: 12.7 mm3; B: 15.2 mm3; and C: 17.7 mm3), were printed in triplicate and subjected to four experimental procedures. Eggs from a housefly colony were suspended in water, sucked up with the 3D-printed tip mounted on a Pasteur plastic pipette and released on a piece of coffee filter. Repeatability intra-run and reproducibility inter-day were evaluated by counting the eggs manipulated with the experimental tips. Time needed for egg sowing via a pipette tip was compared to time needed for counting 100 eggs delivered by a Paster pipette (control). Potential damage to eggs due to the pipette tip was estimated by quantifying fecundity (egg eclosion and larval survival). Coefficients of variation intra-run and inter-day were similar over replicates. Pipette set B had the best repeatability (coefficient of variation intra-run between 4.4 and 4.7%), and pipette set C the highest reproducibility (coefficient of variation inter-day of 2.6%). Egg damage due to the tip was negligible, however the tips did reduce larval survival (between 46.1±7.2% and 63.4±18.9%), compared to the control (between 66.0±2.4 and 75.5±9.3%). The pipette size and number of dispensed eggs correlated well (R2=0.90,P&lt;0.001), but the relation was non-linear (y = 0.051034 x2+ 12.356 x – 0.03627). The time needed to sow a fixed number of eggs on the rearing substrate was reduced by 73% (pipette A) – 80% (pipette C) by using the pipette tip, compared to visual quantification.

A crucial phase in plants - it's a gas, gas, gas!

A crucial phase in plants - it's a gas, gas, gas!