Labeling Of Blood Cells Research Articles

Developments in Transfusion Medicine: Pulmonary Transfusion Reactions and Novel Blood Cell Labeling Techniques.

Staying updated on advancements in transfusion medicine is crucial, especially in critical care and perioperative setting, where timely and accurate transfusions can be lifesaving therapeutic interventions. This narrative review explores the landscape of transfusion-related adverse events, focusing on pulmonary transfusion reactions such as transfusion-associated circulatory overload (TACO) and transfusion-related acute lung injury (TRALI). TACO and TRALI are the leading causes of transfusion-related morbidity and mortality; however, specific treatments are lacking. Understanding the current incidence, diagnostic criteria, pathogenesis, treatment, and prevention strategies can equip clinicians to help reduce the incidence of these life-threatening complications. The review discusses emerging pathogenic mechanisms, including the possible role of inflammation in TACO and the mechanisms of reverse TRALI and therapeutic targets for TACO and TRALI, emphasizing the need for further research to uncover preventive and treatment modalities. Despite advancements, significant gaps remain in our understanding of what occurs during transfusions, highlighting the necessity for improved monitoring methods. To address this, the review also presents novel blood cell labeling techniques in transfusion medicine used for improving monitoring, quality assessment, and as a consequence, potentially reducing transfusion-related complications. This article aims to provide an update for anesthesiologists, critical care specialists, and transfusion medicine professionals regarding recent advancements and developments in the field of transfusion medicine.

Classifying Anemia Diseases based on VGG-16 and CBAM Attention Mechanism Models

Anemia is a major public health concern and has a significant impact on women and children worldwide. This condition is caused by a lack of sufficient Red Blood Cells (RBCs). Traditionally, anemia diagnosis has relied on invasive techniques that require blood samples, leading to pain and discomfort. This study explores deep learning techniques for automating and enhancing anemia detection accuracy using the Blood Cell Count and Detection (BCCD) dataset. We employ the VGG16 convolutional neural network architecture augmented with the Convolutional Block Attention Module (CBAM) to boost feature representation and performance. The BCCD dataset, consisting of 364 images with 4888 labeled blood cells, was used for training and evaluation. Our enhanced VGG16 model achieved accuracy for RBCs, WBCs, and platelets with values of 0.84, 0.93, and 0.92, respectively, effectively identifying various blood cell types and detecting anemia. Precision-recall analysis and confusion matrix metrics confirmed the robustness of the model, with high precision and recall rates and minimal false positives and negatives. These results suggest that advanced deep learning models with attention mechanisms, such as CBAM, can significantly improve medical diagnostics by providing reliable, efficient, and accurate early disease detection tools. Future development will be directed towards fine-tuning and validating the proposed model on other medical imaging datasets for clinical applications.

Structured adaptive boosting trees for detection of multicellular aggregates in fluorescence intravital microscopy

Fluorescence intravital microscopy captures large data sets of dynamic multicellular interactions within various organs such as the lungs, liver, and brain of living subjects. In medical imaging, edge detection is used to accurately identify and delineate important structures and boundaries inside the images. To improve edge sharpness, edge detection frequently requires the inclusion of low-level features. Herein, a machine learning approach is needed to automate the edge detection of multicellular aggregates of distinctly labeled blood cells within the microcirculation. In this work, the Structured Adaptive Boosting Trees algorithm (AdaBoost.S) is proposed as a contribution to overcome some of the edge detection challenges related to medical images. Algorithm design is based on the observation that edges over an image mask often exhibit special structures and are interdependent. Such structures can be predicted using the features extracted from a bigger image patch that covers the image edge mask. The proposed AdaBoost.S is applied to detect multicellular aggregates within blood vessels from the fluorescence lung intravital images of mice exposed to e-cigarette vapor. The predictive capabilities of this approach for detecting platelet-neutrophil aggregates within the lung blood vessels are evaluated against three conventional machine learning algorithms: Random Forest, XGBoost and Decision Tree. AdaBoost.S exhibits a mean recall, F-score, and precision of 0.81, 0.79, and 0.78, respectively. Compared to all three existing algorithms, AdaBoost.S has statistically better performance for recall and F-score. Although AdaBoost.S does not outperform Random Forest in precision, it remains superior to the XGBoost and Decision Tree algorithms. The proposed AdaBoost.S is widely applicable to analysis of other fluorescence intravital microscopy applications including cancer, infection, and cardiovascular disease.

Detection and clinical application of red blood cell survival.

There are 2 techniques for detecting red blood cell survival (RBCS) detection techniques: red blood cell labeling test and carbon monoxide (CO) breath test. The former has disadvantages such as long measurement times and complicated procedures, while the latter is simple, convenient, moderately priced, and capable of dynamically monitoring changes in RBCS before and after treatment. Currently, the CO breath test is gradually being implemented in clinical practice. RBCS is not only applied to hematologic diseases such as multiple myeloma, myelodysplastic syndromes, lymphoma, and thalassemia, but also to non-hematologic diseases like type 2 diabetes and chronic kidney disease. It can assist in diagnosis, guide treatment, evaluate drug treatment efficacy, and predict disease progression.

DeepIFC: Virtual fluorescent labeling of blood cells in imaging flow cytometry data with deep learning.

Imaging flow cytometry (IFC) combines flow cytometry with microscopy, allowing rapid characterization of cellular and molecular properties via high-throughput single-cell fluorescent imaging. However, fluorescent labeling is costly and time-consuming. We present a computational method called DeepIFC based on the Inception U-Net neural network architecture, able to generate fluorescent marker images and learn morphological features from IFC brightfield and darkfield images. Furthermore, the DeepIFC workflow identifies cell types from the generated fluorescent images and visualizes the single-cell features generated in a 2D space. We demonstrate that rarer cell types are predicted well when a balanced data set is used to train the model, and the model is able to recognize red blood cells not seen during model training as a distinct entity. In summary, DeepIFC allows accurate cell reconstruction, typing and recognition of unseen cell types from brightfield and darkfield images via virtual fluorescent labeling.

Magnetic Platelets as a Platform for Drug Delivery and Cell Trapping.

The possibility of using magnetically labeled blood cells as carriers is a novel approach in targeted drug-delivery systems, potentially allowing for improved bloodstream delivery strategies. Blood cells already meet the requirements of biocompatibility, safety from clotting and blockage of small vessels. It would solve the important problem of the patient's immune response to embedded foreign carriers. The high efficiency of platelet loading makes them promising research objects for the development of personalized drug-delivery systems. We are developing a new approach to use platelets decorated with magnetic nanoparticles as a targeted drug-delivery system, with a focus on bloodstream delivery. Platelets are non-nuclear blood cells and are of great importance in the pathogenesis of blood-clotting disorders. In addition, platelets are able to attach to circulating tumor cells. In this article, we studied the effect of platelets labeled with BSA-modified magnetic nanoparticles on healthy and cancer cells. This opens up broad prospects for future research based on the delivery of specific active substances by this method.

Evaluation of different 89Zr-labeled synthons for direct labeling and tracking of white blood cells and stem cells in healthy athymic mice

Cell based therapies are evolving as an effective new approach to treat various diseases. To understand the safety, efficacy, and mechanism of action of cell-based therapies, it is imperative to follow their biodistribution noninvasively. Positron-emission-tomography (PET)-based non-invasive imaging of cell trafficking offers such a potential. Herein, we evaluated and compared three different ready-to-use direct cell radiolabeling synthons, [89Zr]Zr-DFO-Bn-NCS, [89Zr]Zr-Hy3ADA5-NCS, and [89Zr]Zr-Hy3ADA5-SA for PET imaging-based trafficking of white blood cells (WBCs) and stem cells (SCs) up to 7 days in athymic nude mice. We compared the degree of 89Zr complexation and percentage of cell radiolabeling efficiencies with each. All three synthons, [89Zr]Zr-DFO-Bn-NCS, [89Zr]Zr-Hy3ADA5-NCS, and [89Zr]Zr-Hy3ADA5-SA, were successfully prepared, and used for radiolabeling of WBCs and SCs. The highest cell radiolabeling yield was found for [89Zr]Zr-DFO-Bn-NCS, followed by [89Zr]Zr-Hy3ADA5-NCS, and [89Zr]Zr-Hy3ADA5-SA. In terms of biodistribution, WBCs radiolabeled with [89Zr]Zr-DFO-Bn-NCS or [89Zr]Zr-Hy3ADA5-NCS, were primarily accumulated in liver and spleen, whereas SCs radiolabeled with [89Zr]Zr-DFO-Bn-NCS or [89Zr]Zr-Hy3ADA5-NCS were found in lung, liver and spleen. A high bone uptake was observed for both WBCs and SCs radiolabeled with [89Zr]Zr-Hy3ADA5-SA, suggesting in-vivo instability of [89Zr]Zr-Hy3ADA5-SA synthon. This study offers an appropriate selection of ready-to-use radiolabeling synthons for noninvasive trafficking of WBCs, SCs and other cell-based therapies.

The oral ferroportin inhibitor vamifeport improves hemodynamics in a mouse model of sickle cell disease

AbstractSickle cell disease (SCD) is an inherited hemolytic anemia caused by a single point mutation in the β-globin gene of hemoglobin that leads to synthesis of sickle hemoglobin (HbS) in red blood cells (RBCs). HbS polymerizes in hypoxic conditions, leading to intravascular hemolysis, release of free hemoglobin and heme, and increased adhesion of blood cells to the endothelial vasculature, which causes painful vaso-occlusion and organ damage. HbS polymerization kinetics are strongly dependent on the intracellular HbS concentration; a relatively small reduction in cellular HbS concentration may prevent HbS polymerization and its sequelae. We hypothesized that iron restriction via blocking ferroportin, the unique iron transporter in mammals, might reduce HbS concentration in RBCs, thereby decreasing hemolysis, improving blood flow, and preventing vaso-occlusive events. Indeed, vamifeport (also known as VIT-2763), a clinical-stage oral ferroportin inhibitor, reduced hemolysis markers in the Townes model of SCD. The RBC indices of vamifeport-treated male and female Townes mice exhibited changes attributable to iron-restricted erythropoiesis: decreased corpuscular hemoglobin concentration mean and mean corpuscular volume, as well as increased hypochromic and microcytic RBC fractions. Furthermore, vamifeport reduced plasma soluble VCAM-1 concentrations, which suggests lowered vascular inflammation. Accordingly, intravital video microscopy of fluorescently labeled blood cells in the microvasculature of Townes mice treated with vamifeport revealed diminished adhesion to the endothelium and improved hemodynamics. These preclinical data provide a strong proof-of-concept for vamifeport in the Townes model of SCD and support further development of this compound as a potential novel therapy in SCD.

Paper-Based Cytometer for the Detection and Enumeration of White Blood Cells According to Their Immunophenotype.

Total and differential white blood cell (WBC) counts are vital metrics used routinely by clinicians to aid in the identification of diseases. However, the equipment necessary to perform WBC counts restricts their operation to centralized laboratories, greatly limiting their accessibility. Established solutions for the development of point-of-care assays, namely lateral flow tests and paper-based microfluidic devices, are inherently limited in their ability to support the detection of WBCs─the pore sizes of materials used to fabricate these devices (e.g., membranes or chromatography papers) do not permit passive WBC transport via wicking. Herein, we identify a material capable of the unimpeded transport of WBCs in both lateral and vertical directions: a coffee filter. Through in situ labeling with an enzyme-labeled affinity reagent, our paper-based cytometer detects WBCs according to their immunophenotype. Using two cultured leukocyte lines (Jurkat D1.1 T cells and MAVER-1 B cells), we demonstrate the specific, colorimetric enumeration of each target cell population across the expected physiological range for total lymphocytes, 1000-4000 cells μL-1. Additionally, we highlight a potential application of this type of device as a screening tool for detecting abnormal cell counts outside the normal physiological range and in subclasses of cell types, which could aid in the identification of certain diseases (e.g., CD4+ T lymphocytes, an important biomarker for HIV disease/AIDS). These results pave the way for a new class of paper-based devices─those capable of controlled white blood cell transport, labeling, capture, and detection─thus expanding the opportunities for low-cost, point-of-care cytometers.

Development of Mechanical Stability in Late-Stage Embryonic Erythroid Cells: Insights From Fluorescence Imaged Micro-Deformation Studies.

The combined use of fluorescence labeling and micro-manipulation of red blood cells has proven to be a powerful tool for understanding and characterizing fundamental mechanisms underlying the mechanical behavior of cells. Here we used this approach to study the development of the membrane-associated cytoskeleton (MAS) in primary embryonic erythroid cells. Erythropoiesis comes in two forms in the mammalian embryo, primitive and definitive, characterized by intra- and extra-vascular maturation, respectively. Primitive erythroid precursors in the murine embryo first begin to circulate at embryonic day (E) 8.25 and mature as a semi-synchronous cohort before enucleating between E12.5 and E16.5. Previously, we determined that the major components of the MAS become localized to the membrane between E10.5 and E12.5, and that this localization is associated with an increase in membrane mechanical stability over this same period. The change in mechanical stability was reflected in the creation of MAS-free regions of the membrane at the tips of the projections formed when cells were aspirated into micropipettes. The tendency to form MAS-free regions decreases as primitive erythroid cells continue to mature through E14.5, at least 2 days after all detectable cytoskeletal components are localized to the membrane, indicating continued strengthening of membrane cohesion after membrane localization of cytoskeletal components. Here we demonstrate that the formation of MAS-free regions is the result of a mechanical failure within the MAS, and not the detachment of membrane bilayer from the MAS. Once a “hole” is formed in the MAS, the skeletal network contracts laterally along the aspirated projection to form the MAS-free region. In protein 4.1-null primitive erythroid cells, the tendency to form MAS-free regions is markedly enhanced. Of note, similar MAS-free regions were observed in maturing erythroid cells from human marrow, indicating that similar processes occur in definitive erythroid cells. We conclude that localization of cytoskeletal components to the cell membrane of mammalian erythroid cells during maturation is insufficient by itself to produce a mature MAS, but that subsequent processes are additionally required to strengthen intraskeletal interactions.

UHAS-MIDA Software Package: Mass Isotopomer Distribution Analysis-Applying the Inverse Matrix for the Determination of Stable Isotopes of Chromium Distribution during Red Blood Cell Labelling

Clinical assessment of fluid volume status in children during malaria can be taxing and often inaccurate. During malaria, changes in fluid volume are rather multifarious and estimating this parameter, especially in sick children is very challenging for clinicians who frequently rely on indices such as long capillary refill times, tachycardia, central venous pressure and decreased urine volume as guides. Here, we present the UHAS-MIDA, an open-source software tool that calculates the red blood cell (RBC) concentration and blood volume during malaria in children determined using a stable isotope of chromium (53Cr as the label) by gas chromatography-mass spectrometry in selective ion monitoring (GC/MS-SIM) analysis. A key component involves the determination of the compositions of the most abundant naturally occurring isotopes of Cr (50Cr, 52Cr, 53Cr), and converting the proportions into a 3 × 3 matrix. To estimate unknown proportions of chromium isotopic mixtures from the measured abundances of three ions, an inverse matrix was calculated. The inverse together with several inputs is then used to calculate the corrected MS ion abundances. Thus, we constructed the software tool UHAS- MIDA using HTML, CSS/Bootstrap, JavaScript, and PHP scripting languages. The tool enables the user to efficiently determine RBC concentration and fluid volume. The source code, binary packages and associated materials for UHAS-MIDA are freely available at https://github.com/bentil078/Abaye-et-al_UHASmida

Using Flow Cytometry to Detect and Quantitate Altered Blood Formation in the Developing Zebrafish.

The diversity of cell lineages that comprise mature blood in vertebrate animals arise from the differentiation of hematopoietic stem and progenitor cells (HSPCs). This is a critical process that occurs throughout the lifespan of organisms, and disruption of the molecular pathways involved during embryogenesis can have catastrophic long-term consequences. For a multitude of reasons, zebrafish (Danio rerio) has become a model organism to study hematopoiesis. Zebrafish embryos develop externally, and by 7 days postfertilization (dpf) have produced most of the subtypes of definitive blood cells that will persist for their lifetime. Assays to assess the number of hematopoietic cells have been developed, mainly utilizing specific histological stains, in situ hybridization techniques, and microscopy of transgenic animals that utilize blood cell-specific promoters driving the expression of fluorescent proteins. However, most staining assays and in situ hybridization techniques do not accurately quantitate the number of blood cells present; only large differences in cell numbers are easily visualized. Utilizing transgenic animals and analyzing individuals with fluorescent or confocal microscopy can be performed, but the quantitation of these assays relies on either counting manually or utilizing expensive imaging software, both of which can make errors. Development of additional methods to assess blood cell numbers would be economical, faster, and could even be automated to quickly assess the effect of CRISPR-mediated genetic modification, morpholino-mediated transcript reduction, and the effect of drug compounds that affect hematopoiesis on a large scale. This novel assay to quantitate blood cells is performed by dissociating whole zebrafish embryos and analyzing the amount of fluorescently labelled blood cells present. These assays should allow elucidation of molecular pathways responsible for blood cell generation, expansion, and regulation during embryogenesis, which will allow researchers to further discover novel factors altered during blood diseases, as well as pathways essential during the evolution of vertebrate hematopoiesis.

In vivo analysis of noise dependent activation of white blood cells and microvascular dysfunction in mice

This article contains supporting information on data collection for the research article entitled “Aircraft noise exposure drives the activation of white blood cells and induces microvascular dysfunction in mice” by Eckrich et al. We found that noise-induced stress triggered microvascular dysfunction via involvement of innate immune-derived reactive oxygen species. In this article, we present the instrumentation of mice with dorsal skinfold chambers for in vivo microscopic imaging of blood flow, interaction of leukocytes with the vascular wall (also by fluorescent labelling of blood cells) and vessel diameter. In addition, we explain the preparation of cerebral arterioles for measurement of vascular reactivity in vitro.•visualization of noise-dependent effects in dorsal skinfold chamber.•in vivo microscopy of noise-dependent activation of white blood cells.•analysis of noise-dependent microvascular dysfunction in dorsal skinfold chamber and cannulated cerebral arterioles.

Deep Learning Approach for Quantification of Fluorescently Labeled Blood Cells in Danio rerio (Zebrafish).

Neutrophils are a type of white blood cell essential for the function of the innate immune system. To elucidate mechanisms of neutrophil biology, many studies are performed in vertebrate animal model systems. In Danio rerio (zebrafish), in vivo imaging of neutrophils is possible due to transgenic strains that possess fluorescently labeled leukocytes. However, due to the relative abundance of neutrophils, the counting process is laborious and subjective. In this article, we propose the use of a custom trained “you only look once” (YOLO) machine learning algorithm to automate the identification and counting of fluorescently labeled neutrophils in zebrafish. Using this model, we found the correlation coefficient between human counting and the model equals r = 0.8207 with an 8.65% percent error, while variation among human counters was 5% to 12%. Importantly, the model was able to correctly validate results of a previously published article that quantitated neutrophils manually. While the accuracy can be further improved, this model notably achieves these results in mere minutes compared with hours via standard manual counting protocols and can be performed by anyone with basic programming knowledge. It further supports the use of deep learning models for high throughput analysis of fluorescently labeled blood cells in the zebrafish model system.

Detection of Melanoma Cells in Whole Blood Samples Using Spectral Imaging and Optical Clearing

Most cancer deaths are associated with metastases resulting from the spread of circulating tumor cells (CTCs) from the primary tumor to vital organs. The existing methods for detection of CTCs as markers of metastasis progression are time consuming with several steps of sample processing, including red blood cell removal, labeling, immunomagnetic capture and isolation, which can lead to loss of CTCs. Here we introduce a method for detection and identification of CTCs using spectral absorption imaging of melanoma cells and optical clearing of whole blood samples. Verification of this approach was performed using phantoms of human melanoma cells and suspensions of mouse melanoma cells of line B16F10 alone and in mixture with blood. A method for improving detection sensitivity has been demonstrated applying optical clearing of mouse blood using biocompatible chemical agents. The findings suggest that the proposed diagnostic platform has the potential to detect quickly CTCs in whole blood samples from patients with melanoma.

Chromosome aberrations during in-vitro labeling of red blood cells with 99mTc: a study with cytokinesis block micronucleus assay in human peripheral blood lymphocytes.

Radiopharmaceuticals are special medicines composed by a radionuclide and a non-radioactive compound characterized by non-pharmacodynamic effects, low prevalence of side effects, and a possible risk of oncogenesis, since its administration to patients supposes a radiation dose to organism. Over these years, radiation damage induced by diagnosis radiopharmaceuticals has been evaluated, including the radiolabeled autologous cells, a group of radiopharmaceuticals where blood cells extracted from patients are labeled in-vitro and readministered for diagnosis. There is not a consensus about the possibility of increasement of risk for malignancies associated with the radiolabeled blood cells, so for a more accurate evaluation of the potential oncogenic risk related to the administration of [Tc]Tc labeled red blood cells, radiation dose received by the cells during the labeling process is studied by means of the cytokinesis-blocked micronucleus (CBMN) assay and a dose-response curve constructed by in-vitro external irradiation of blood samples. Our work enables to establish the range of activity to be added during the in-vitro labeling of red blood cells with [Tc]Tc pertechnetate to avoid radiation damage to cells. Activities recommended for blood volume determination and angiography do not increase the risk of malignancies, whilst activities of 370 MBq show chromosome aberrations in lymphocytes. Evaluation of the radiation damage related to the in-vitro labeling is recommended to estimate the potential oncogenic risk and minimize it.

A kit formulation for the preparation of [89Zr]Zr(oxinate)4 for PET cell tracking: White blood cell labelling and comparison with [111In]In(oxinate)3

BackgroundAdvances in immunology and cell-based therapies are creating a need to track individual cell types, such as immune cells (neutrophils, eosinophils, chimeric antigen receptor (CAR) T cells, etc.) and stem cells. As the fate of administered cells remains largely unknown, nuclear imaging could determine the migration and survival of cells in patients. [89Zr]Zr(oxinate)4, or [89Zr]Zr-oxine, is a radiotracer for positron emission tomography (PET) that has been evaluated in preclinical models of cell tracking and could improve on [111In]In-oxine, the current gold standard radiotracer for cell tracking by scintigraphy and single-photon emission computed tomography (SPECT), because of the better sensitivity, spatial resolution and quantification of PET. However, a clinically usable formulation of [89Zr]Zr-oxine is lacking. This study demonstrates a 1-step procedure for preparing [89Zr]Zr-oxine and evaluates it against [111In]In-oxine in white blood cell (WBC) labelling. MethodsCommercial [89Zr]Zr-oxalate was added to a formulation containing oxine, a buffering agent, a base and a surfactant or organic solvent. WBC isolated from 10 human volunteers were radiolabelled with [89Zr]Zr-oxine following a clinical radiolabelling protocol. Labelling efficiency, cell viability, chemotaxis and DNA damage were evaluated in vitro, in an intra-individual comparison against [111In]In-oxine. ResultsAn optimised formulation of [89Zr]Zr-oxine containing oxine, polysorbate 80 and 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) was developed. This enabled 1-step radiolabelling of oxine with commercial [89Zr]Zr-oxalate (0.1–25 MBq) in 5 min and radiotracer stability for 1 week. WBC labelling efficiency was 48.7 ± 6.3%, compared to 89.1 ± 9.5% (P < 0.0001, n = 10) for [111In]In-oxine. Intracellular retention of 89Zr and cell viability after radiolabelling were comparable to 111In. There were no significant differences in leukocyte chemotaxis or DNA damage between [89Zr]Zr-oxine or [111In]In-oxine. Conclusions, advances in knowledge and implications for patient careOur results demonstrate that [89Zr]Zr-oxine is a suitable PET alternative to [111In]In-oxine for WBC imaging. Our formulation allows rapid, stable, high-yield, single-step preparation of [89Zr]Zr-oxine from commercially available 89Zr. This will facilitate the clinical translation of cell tracking using [89Zr]Zr-oxine.

Left ventricular ejection fraction determined with the simulation of a very low-dose CZT-SPECT protocol and an additional count-calibration on planar radionuclide angiographic data

PurposeTo determine whether the left ventricular ejection fractions (EFs), measured on a high-sensitivity CZT single photon emission computed tomography (SPECT)-camera with a 70% reduction in recording times and a prevention of EF overestimation through an additional count-calibration, are concordant with reference EF from planar radionuclide angiography (2D-RNA). MethodsAn additional 10-minute CZT-SPECT recording was performed in patients referred to 2D-RNA for cardiomyopathy (n = 23) or chemotherapy monitoring (n = 50) with an in vivo red blood cell labeling with 850 MBq 99mTcO4-. The EF, obtained from CZT-SPECT with 100% (SPECT100) or 30% (SPECT30) projection times and with a SPECT-count calibration on the 2D-RNA counts of corresponding cavity volumes, were compared to EF from 2D-RNA. ResultsStrong and equivalent relationships were documented between the EF from 2D-RNA and the calibrated EF from SPECT100 (y = 0.89x + 6.62; R2 = 0.87) and SPECT30 (y = 0.87x + 8.40; R2 = 0.85), and the mean EF from SPECT100 (54% ± 15%) and SPECT30 (53% ± 16%) were close to that from 2D-RNA (55% ± 15%). However, upward shifts in these mean values were documented in the absence of count calibration for both SPECT100 (60% ± 18%) and SPECT30 (60% ± 18%). ConclusionLeft ventricular EF may be determined on a high-sensitivity CZT-camera, a 70% reduction in injected activities, and an additional count-calibration for further enhancing the concordance with 2D-RNA values.

Development of Blood-Cell-Selective Fluorescent Biodots for Lysis-Free Leukocyte Imaging and Differential Counting in Whole Blood.

Complete blood count with leukocyte (white blood cell, WBC) differential is one of the most frequently ordered clinical test for disease diagnosis. Herein, multifunctional fluorescent carbon dots derived from biomolecules (biodots) for rapid lysis-free whole blood analysis are developed. Specifically, two types of biodots are molecularly engineered through hydrothermal synthesis for differential blood cells labeling. Type I biodots synthesized from amino acid (serine/threonine) precursors and passivated with polyethylenimine can label both red blood cells (RBCs) and WBCs with excellent contrast in fluorescence intensity, enabling direct counting of leukocytes in whole blood samples without a tedious RBC lysis step. It also allows three-part leukocyte differential counting by flow cytometry without using expensive fluorophore-conjugated antibodies. On the other hand, Type II biodots synthesized from the same amino acid precursors but passivated with a biopolymer (chitosan) are able to selectively lyse RBCs with greater than 98% efficiency to allow simultaneous fluorescent labeling of leukocytes for WBC counting in whole blood. It is envisioned that these novel nanoreagents, which eliminate the cumbersome lysis and antibody conjugation steps for selective labeling of different blood cells, would revolutionize disease diagnostics toward achieving faster, cheaper, and more accurate whole blood analyses in one test.

A Subpopulation of Monocytes in Normal Human Blood Has Significant Magnetic Susceptibility: Quantification and Potential Implications.

The presence of iron in circulating monocytes is well known as they play essential roles in iron recycling. Also, the storage of this metal as well as its incorrect uptake and/or release are important data to diagnose different pathologies. It has been demonstrated that iron storage in human blood cells can be measured through their magnetic behavior with high accuracy; however, the magnetic characteristics of monocytes have not been reported so far to the best of our knowledge. Therefore, in this work, we report, for the first time, the physical and magnetic properties of human monocytes, along with plasma platelets, oxyhemoglobin red blood cells (oxyHb-RBCs), and methemoglobin red blood cells (metHb-RBCs). The different cell populations were separated by Ficoll-density gradient centrifugation, followed by a flow sorting step to isolate monocytes from peripheral blood mononuclear cells. The different fractions were analyzed by Coulter Counter (for determining the size distribution and concentration) and the sorted monocytes were qualitatively analyzed on ImageStream, a state-of-the-art imaging cytometer. The analysis of the Coulter Counter and ImageStream data suggests that although there exists contamination in the monocyte fraction, the integrity of the sorted monocytes appears to be intact and the concentration was high enough to precisely measure their magnetic velocity by Cell Tracking Velocimetry. Surprisingly, monocytes reported the highest magnetic mobility from the four fractions under analysis, with an average magnetic velocity 7.8 times higher than MetHb-RBCs, which is the only type of cells with positive magnetic velocities. This value is equivalent to a susceptibility 2.5 times higher than the value reported by fresh MetHb-RBCs. It should be noted that this is the first study that reports that a subpopulation of human monocytes is much more magnetic than MetHb-RBCs, opening the door to the possible isolation of human monocytes by label-free magnetic techniques. Further, it is suggested that these magnetic monocytes could "contaminate" positively selected, immunomagnetically labeled blood cells (i.e., during a process using magnetically conjugated antibodies targeting cells, such as CD34 positive cells). Conversely, these magnetic monocytes could be inadvertently removed from a desired blood population when one is using a negative magnetic isolation technique to target cells for removal. © 2019 International Society for Advancement of Cytometry.