Clinical laboratory test utilization of CSF oligoclonal bands and IgG index in a tertiary pediatric hospital.

BACKGROUND: Criteria developed for the diagnosis of multiple sclerosis (MS) in adults are also used in the pediatric setting. However, differential diagnosis in pediatric-onset MS (POMS) is distinct from that of adult-onset MS. There is little literature characterizing the utility of oligoclonal bands (OCB) and IgG index in differentiating POMS from other childhood diseases with overlapping clinical presentation which can require immediate treatment. METHODS: A retrospective review of all MS panels resulted between March 2022 and May 2023 on patients age ≤ 18 years at one tertiary care pediatric hospital in the northeastern United States was performed with pediatric neurology collaboration to characterize clinical utility (n = 85 cases). RESULTS: Demyelinating diseases accounted for 31 of 85 total cases (36.5%), 12 of these cases were POMS (14%). Other diagnoses consisted of psychiatric etiologies (17.6%), infectious meningitis/encephalitis (5.9%), and migraine (5.9%). Elevated IgG index was seen in 67% of those with demyelinating diseases, versus only 13% of those with other conditions. Unique OCBs were found in 41% of those with demyelinating diseases, versus only 9% of those with other conditions. Fourteen of 15 patients (93.3%) with psychiatric conditions had normal MS panels. CONCLUSIONS: Patients with demyelinating diseases were more likely to have elevated IgG index and unique OCBs versus patients with other conditions. For pediatric hospitals without in-house OCB evaluation, implementation of an in-house IgG index may serve as a rapid screen for differentials that include demyelinating diseases while awaiting OCB results, in the appropriate clinical context. IMPACT STATEMENT: IgG index and CSF oligoclonal bands are important tools in the diagnosis of patients with suspected Multiple Sclerosis (MS). In the pediatric population, these markers are used to differentiate pediatric-onset MS (POMS) from other neurologic, psychiatric, and inflammatory diseases that display clinical overlap. The use of these markers in differentiating these conditions has not been thoroughly investigated. We examined the associations between abnormal markers and final diagnoses in pediatric patients undergoing testing for POMS in order to identify trends that may enhance ordering and reporting practices.

Radiomodulating Properties of Superparamagnetic Iron Oxide Nanoparticle (SPION) Agent Ferumoxytol on Human Monocytes: Implications for MRI-Guided Liver Radiotherapy.

Superparamagnetic iron oxide nanoparticles (SPION) have attracted great attention not only for therapeutic applications but also as an alternative magnetic resonance imaging (MRI) contrast agent that helps visualize liver tumors during MRI-guided stereotactic body radiotherapy (SBRT). SPION can provide functional imaging of liver parenchyma based upon its uptake by the hepatic resident macrophages or Kupffer cells with a relative enhancement of malignant tumors that lack Kupffer cells. However, the radiomodulating properties of SPION on liver macrophages are not known. Utilizing human monocytic THP-1 undifferentiated and differentiated cells, we characterized the effect of ferumoxytol (Feraheme®), a carbohydrate-coated ultrasmall SPION agent at clinically relevant concentration and therapeutically relevant doses of gamma radiation on cultured cells in vitro. We showed that ferumoxytol affected both monocytes and macrophages, increased the resistance of monocytes to radiation-induced cell death and inhibition of cell activity, and supported the anti-inflammatory phenotype of human macrophages under radiation. Its effect on human cells depended on the duration of SPION uptake and was radiation dose-dependent. The results of this pilot study support a strong mechanism-based optimization of SPION-enhanced MRI-guided liver SBRT for primary and metastatic liver tumors, especially in patients with liver cirrhosis awaiting a liver transplant.

Clinical Significance of Uncommon, Non-Clinical, and Novel Autoantibodies.

Autoantibodies are a common mark of autoimmune reaction and their identification in the patients' serum, cerebrospinal fluid, or tissues is generally believed to represent diagnostic or prognostic biomarkers of autoimmune diseases or autoinflammatory conditions. Traditionally, autoantibody testing is an important part of the clinical examination of suspected patients, and in the absence of reliable T cell tests, characterization of autoantibody responses might be suitable in finding causes of specific autoimmune responses, their strength, and sometimes commencement of autoimmune disease. Autoantibodies are also useful for prognostic stratification in clinically diverse groups of patients if checked repeatedly. Antibody discoveries are continuing, with important consequences for verifying autoimmune mechanisms, diagnostic feasibility, and clinical management. Adding newly identified autoantibody-autoantigen pairs to common clinical laboratory panels should help upgrade and harmonize the identification of systemic autoimmune rheumatic disorders and other autoimmune conditions. Herein, we aim to summarize our current knowledge of uncommon and novel autoantibodies in the context of discussing their validation, diagnostic practicability, and clinical relevance. The regular updates within the field are important and well justified.

Schwann cells in the normal and pathological lung microenvironment.

The lungs are a key organ in the respiratory system. They are regulated by a complex network of nerves that control their development, structure, function, and response to various pathological stimuli. Accumulating evidence suggests the involvement of a neural mechanism in different pathophysiological conditions in the lungs and the development and progression of common respiratory diseases. Lung diseases are the chief source of death globally. For instance, lung cancer is the second most commonly diagnosed malignancy, after prostate cancer in men and breast cancer in women, and is the most lethal cancer worldwide. However, although airway nerves are accepted as a mechanistically and therapeutically important feature that demands appropriate emphasizing in the context of many respiratory diseases, significantly less is known about the role of the neuroglial cells in lung physiology and pathophysiology, including lung cancer. New data have uncovered some cellular and molecular mechanisms of how Schwann cells, as fundamental components of the peripheral nervous system, may regulate lung cancer cells' survival, spreading, and invasiveness in vitro and in vivo. Schwann cells control the formation and maintenance of the lung cancer microenvironment and support metastasis formation. It was also reported that the number of lung cancer-associated Schwann cells correlates with patients' survival. Different factors secreted by Schwann cells, including microRNA, are known to sharpen the lung cancer environment by regulating the tumor-neuro-immune axis. Further clinical and experimental studies are required to elucidate the detailed role of Schwann cells in creating and maintaining pulmonary tumor-neuro-immune axis, which will advance our understanding of the pathogenesis of lung cancer and may inform therapeutic hypotheses aiming neoplasms and metastases in the lung.

Diagnostics of Tuberculosis with Single-Walled Carbon Nanotube-Based Field-Effect Transistors.

Tuberculosis (TB) is still threatening millions of people's lives, especially in developing countries. One of the major factors contributing to the ongoing epidemic of TB is the lack of a fast, efficient, and inexpensive diagnostic strategy. In this work, we developed a semiconducting single-walled carbon nanotube (SWCNT)-based field-effect transistor (FET) device functionalized with anti-Mycobacterium tuberculosis antigen 85B antibody (Ab85B) to detect the major M. tuberculosis-secreted antigen 85B (Ag85B). Through optimizing the device fabrication process by evaluating the mass of the antibody and the concentration of the gating electrolyte, our Ab85B-SWCNT FET devices achieved the detection of the Ag85B spiked in phosphate-buffered saline (calibration samples) with a limit of detection (LOD) of 0.05 fg/mL. This SWCNT FET biosensor also showed good sensing performance in biological matrices including artificial sputum and can identify Ag85B in serum after introducing bovine serum albumin (BSA) into the blocking layer. Furthermore, our BSA-blocked Ab85B-SWCNT FET devices can distinguish between TB-positive and -negative clinical samples, promising the application of SWCNT FET devices in point-of-care TB diagnostics. Moreover, the robustness of this SWCNT-based biosensor to the TB diagnosis in blood serum was enhanced by blocking SWCNT devices directly with a glutaraldehyde cross-linked BSA layer, enabling future applications of these SWCNT-based biosensors in clinical testing.

Schwann cell-derived exosomes promote lung cancer progression via miRNA-21-5p.

Schwann cells (SCs), the primary glial cells of the peripheral nervous system, which have been identified in many solid tumors, play an important role in cancer development and progression by shaping the tumor immunoenvironment and supporting the development of metastases. Using different cellular, molecular, and genetic approaches with integrated bioinformatics analysis and functional assays, we revealed the role of human SC-derived exosomal miRNAs in lung cancer progression in vitro and in vivo. We found that exosomal miRNA-21 from SCs up-regulated the proliferation, motility, and invasiveness of human lung cancer cells in vitro, which requires functional Rab small GTPases Rab27A and Rab27B in SCs for exosome release. We also revealed that SC exosomal miRNA-21-5p regulated the functional activation of tumor cells by targeting metalloprotease inhibitor RECK in tumor cells. Integrated bioinformatic analyses showed that hsa-miRNA-21-5p is associated with poor prognosis in patients with lung adenocarcinoma and can promote lung cancer progression through multiple signaling pathways including the MAPK, PI3K/Akt, and TNF signaling. Furthermore, in mouse xenograft models, SC exosomes and SC exosomal hsa-miRNA-21-5p augmented human lung cancer cell growth and lymph node metastasis in vivo. Together our data revealed, for the first time, that SC-secreted exosomes and exosomal miRNA-21-5p promoted the proliferation, motility, and spreading of human lung cancer cells in vitro and in vivo. Thus, exosomal miRNA-21 may play an oncogenic role in SC-accelerated progression of lung cancer and this pathway may serve as a new therapeutic target for further evaluation.

Tixagevimab Plus Cilgavimab Does Not Affect the Interpretation of Electrophoretic and Free Light Chain Assays.

OBJECTIVES: There is concern that the anti-severe acute respiratory syndrome coronavirus 2 therapeutic monoclonal antibodies, used as preexposure prophylaxis in patients with multiple myeloma, may appear as a detectable monoclonal protein by electrophoretic methods, resulting in misinterpretation or inability to measure therapeutic responses in some patients. In this pilot study, we characterize the effect of tixagevimab plus cilgavimab (Evusheld; T + C) on interpretation of serum protein electrophoresis (SPE), immunofixation electrophoresis (IFE), and serum free light chain (sFLC) assays. METHODS: We performed spiking experiments with T + C at serum maximum concentration following a 300-mg dose (1× Cmax) and at 10 times the concentration of Cmax (10× Cmax) with pooled serum samples. SPE and IFE technical procedures were performed on the SPIFE 3000, and sFLC and immunoglobulin G1 (IgG1) subtype quantitation was performed on the Optilite. RESULTS: T + C-associated interference was not visible as an M-spike in normogammaglobulinemic pooled samples. Hypogammaglobulemic pooled samples at 10× Cmax demonstrated an M-spike in SPE and immunoglobulin Gκ pattern in IFE. No increases were noted in the results of sFLC or IgG1 levels. CONCLUSIONS: This study indicates that T + C at pharmacologic Cmax is unlikely to interfere with SPE, IFE, sFLC, or IgG1 analyses when spiked into patient serum samples, but further evaluation of recently injected patients may be warranted.

A Novel Therapeutic Target for Small-Cell Lung Cancer: Tumor-Associated Repair-like Schwann Cells.

Small-cell lung cancer (SCLC), representing 15-20% of all lung cancers, is an aggressive malignancy with a distinct natural history, poor prognosis, and limited treatment options. We have previously identified Schwann cells (SCs), the main glial cells of the peripheral nervous system, in tumor tissues and demonstrated that they may support tumor spreading and metastasis formation in the in vitro and in vivo models. However, the role of SCs in the progression of SCLC has not been investigated. To clarify this issue, the cell proliferation assay, the annexin V apoptosis assay, and the transwell migration and invasion assay were conducted to elucidate the roles in SCLC of tumor-associated SCs (TA-SCs) in the proliferation, apoptosis, migration, and invasion of SCLC cells in vitro, compared to control group. In addition, the animal models to assess SC action's effects on SCLC in vivo were also developed. The result confirmed that TA-SCs have a well-established and significant role in facilitating SCLC cell cancer migration and invasion of SCLC in vitro, and we also observed that SC promotes tumor growth of SCLC in vivo and that TA-SCs exhibited an advantage and show a repair-like phenotype, which allowed defining them as tumor-associated repair SCs (TAR-SCs). Potential molecular mechanisms of pro-tumorigenic activity of TAR-SCs were investigated by the screening of differentially expressed genes and constructing networks of messenger-, micro-, and long- non-coding RNA (mRNA-miRNA-lncRNA) using DMS114 cells, a human SCLC, stimulated with media from DMS114-activated SCs, non-stimulated SCs, and appropriate controls. This study improves our understanding of how SCs, especially tumor-activated SCs, may promote SCLC progression. Our results highlight a new functional phenotype of SCs in cancer and bring new insights into the characterization of the nervous system-tumor crosstalk.

Tumor-Induced T Cell Polarization by Schwann Cells.

Nerve-cancer crosstalk resulting in either tumor neurogenesis or intratumoral neurodegeneration is critically controlled by Schwann cells, the principal glial cells of the peripheral nervous system. Though the direct stimulating effect of Schwann cells on malignant cell proliferation, motility, epithelial-mesenchymal transition, and the formation of metastases have been intensively investigated, the ability of Schwann cells to affect the effector and regulatory immune cells in the tumor environment is significantly less studied. Here, we demonstrated that tumor cells could stimulate Schwann cells to produce high levels of prostaglandin E, which could be blocked by COX-2 inhibitors. This effect was mediated by tumor-derived TGF-ß as neutralization of this cytokine in the tumor-conditioned medium completely blocked the inducible prostaglandin E production by Schwann cells. Similar protective effects were also induced by the Schwann cell pretreatment with TGF-ßR1/ALK4/5/7 and MAPK/ERK kinase inhibitors of the canonical and non-canonical TGF-ß signaling pathways, respectively. Furthermore, prostaglandin E derived from tumor-activated Schwann cells blocked the proliferation of CD3/CD28-activated T cells and upregulated the expression of CD73 and PD-1 on both CD4+ and CD8+ T cells, suggesting T cell polarization to the exhausted phenotype. This new pathway of tumor-induced T cell inhibition via the activation of neuroglial cells represents new evidence of the importance of nerve-cancer crosstalk in controlling tumor development and progression. A better understanding of the tumor-neuro-immune axis supports the development of efficient targets for harnessing this axis and improving the efficacy of cancer therapy.

Sensory Nerves Impede the Formation of Tertiary Lymphoid Structures and Development of Protective Antimelanoma Immune Responses.

Peripheral neurons comprise a critical component of the tumor microenvironment (TME). The role of the autonomic innervation in cancer has been firmly established. However, the effect of the afferent (sensory) neurons on tumor progression remains unclear. Utilizing surgical and chemical skin sensory denervation methods, we showed that afferent neurons supported the growth of melanoma tumors in vivo and demonstrated that sensory innervation limited the activation of effective antitumor immune responses. Specifically, sensory ablation led to improved leukocyte recruitment into tumors, with decreased presence of lymphoid and myeloid immunosuppressive cells and increased activation of T-effector cells within the TME. Cutaneous sensory nerves hindered the maturation of intratumoral high endothelial venules and limited the formation of mature tertiary lymphoid-like structures containing organized clusters of CD4+ T cells and B cells. Denervation further increased T-cell clonality and expanded the B-cell repertoire in the TME. Importantly, CD8a depletion prevented denervation-dependent antitumor effects. Finally, we observed that gene signatures of inflammation and the content of neuron-associated transcripts inversely correlated in human primary cutaneous melanomas, with the latter representing a negative prognostic marker of patient overall survival. Our results suggest that tumor-associated sensory neurons negatively regulate the development of protective antitumor immune responses within the TME, thereby defining a novel target for therapeutic intervention in the melanoma setting.

Cross-talk between HIF and PD-1/PD-L1 pathways in carcinogenesis and therapy.

Tumor-associated hypoxia plays an important role in carcinogenesis and metastasis. The expression, activation, and stabilization of hypoxia-inducible transcription factors (HIFs) support malignant cell survival, proliferation, plasticity, and motility. Hypoxia also upregulates the expression of programmed cell death ligand 1 (PD-L1) in malignant and immune regulatory cells. Therefore, the combination of HIF inhibitors and checkpoint inhibitors (CPIs) is promising for boosting antitumor immunity and diminishing malignant cell plasticity and therapy resistance. In this issue of the JCI, Salman et al. report the development of a specific agent that inhibited HIF-1/2-mediated gene expression in tumor cells and suppressed tumor growth. Bailey, Liu, et al. in this issue demonstrate that targeting HIF-1α abrogated PD-L1-mediated immune evasion by suppressing PD-L1 expression on malignant and myeloid regulatory cells, causing tumor rejection. These findings suggest that targeting the HIF/PD-L1 axis with specific HIF inhibitors should improve the safety and efficacy of CPIs for cancer therapy.

Regulation of Carcinogenesis by Sensory Neurons and Neuromediators.

Interactions between the immune system and the nervous system are crucial in maintaining homeostasis, and disturbances of these neuro-immune interactions may participate in carcinogenesis and metastasis. Nerve endings have been identified within solid tumors in humans and experimental animals. Although the involvement of the efferent sympathetic and parasympathetic innervation in carcinogenesis has been extensively investigated, the role of the afferent sensory neurons and the neuropeptides in tumor development, growth, and progression is recently appreciated. Similarly, current findings point to the significant role of Schwann cells as part of neuro-immune interactions. Hence, in this review, we mainly focus on local and systemic effects of sensory nerve activity as well as Schwann cells in carcinogenesis and metastasis. Specific denervation of vagal sensory nerve fibers, or vagotomy, in animal models, has been reported to markedly increase lung metastases of breast carcinoma as well as pancreatic and gastric tumor growth, with the formation of liver metastases demonstrating the protective role of vagal sensory fibers against cancer. Clinical studies have revealed that patients with gastric ulcers who have undergone a vagotomy have a greater risk of stomach, colorectal, biliary tract, and lung cancers. Protective effects of vagal activity have also been documented by epidemiological studies demonstrating that high vagal activity predicts longer survival rates in patients with colon, non-small cell lung, prostate, and breast cancers. However, several studies have reported that inhibition of sensory neuronal activity reduces the development of solid tumors, including prostate, gastric, pancreatic, head and neck, cervical, ovarian, and skin cancers. These contradictory findings are likely to be due to the post-nerve injury-induced activation of systemic sensory fibers, the level of aggressiveness of the tumor model used, and the local heterogeneity of sensory fibers. As the aggressiveness of the tumor model and the level of the inflammatory response increase, the protective role of sensory nerve fibers is apparent and might be mostly due to systemic alterations in the neuro-immune response. Hence, more insights into inductive and permissive mechanisms, such as systemic, cellular neuro-immunological mechanisms of carcinogenesis and metastasis formation, are needed to understand the role of sensory neurons in tumor growth and spread.

Tumor Innervation: History, Methodologies, and Significance.

The role of the nervous system in cancer development and progression has been under experimental and clinical investigation since nineteenth-century observations in solid tumor anatomy and histology. For the first half of the twentieth century, methodological limitations and opaque mechanistic concepts resulted in ambiguous evidence of tumor innervation. Differential spatial distribution of viable or disintegrated nerve tissue colocalized with neoplastic tissue led investigators to conclude that solid tumors either are or are not innervated. Subsequent work in electrophysiology, immunohistochemistry, pathway enrichment analysis, neuroimmunology, and neuroimmunooncology have bolstered the conclusion that solid tumors are innervated. Regulatory mechanisms for cancer-related neurogenesis, as well as specific operational definitions of perineural invasion and axonogenesis, have helped to explain the consensus observation of nerves at the periphery of the tumor signifying a functional role of nerves, neurons, neurites, and glia in tumor development.

COVID-19 mRNA Vaccines May Cause False Reactivity in Some Serologic Laboratory Tests, Including Rapid Plasma Reagin Tests.

OBJECTIVES: Acute viral infections and some vaccines have been shown to increase false positivity in serologic assays. We assessed if the messenger RNA coronavirus disease 2019 (COVID-19) vaccines could cause false reactivity in common serologic assays in a pilot longitudinal cohort. METHODS: Thirty-eight participants with sera available prevaccination, 2 weeks after each vaccine dose, and monthly thereafter for up to 5 months were tested for common infectious disease serologies and antiphospholipid syndrome (APS) serology markers on the BioPlex 2200, Sure-Vue rapid plasma reagin (RPR), and Macro-Vue RPR. Twenty-two participants received the Moderna vaccine and 16 received the Pfizer vaccine. RESULTS: Most assays had no change in reactivity over the course of the sample draws, including APS markers. Epstein-Barr virus immunoglobulin G (IgG), measles IgG, and rubella immunoglobulin M all had possible false reactivity in one to two participants. RPR tests demonstrated false reactivity, with baseline nonreactive participant samples becoming reactive following vaccination. There were more false reactive participants (7/38) in the BioPlex RPR than in the Sure-Vue (2/38) and Macro-Vue (1/38) tests. All falsely reactive RPR tests were in participants who received the Moderna vaccine. CONCLUSIONS: Serologic assays with results that do not fit the clinical picture following COVID-19 vaccination should be repeated. Effects of false reactivity can last more than 5 months in some assays. In particular, RPR is susceptible to false reactivity, and there is variability among assays. Larger longitudinal studies are needed to determine the incidence and window of false reactivity.

A Carbon Nanotube Sensor Array for the Label-Free Discrimination of Live and Dead Cells with Machine Learning.

Developing robust cell recognition strategies is important in biochemical research, but the lack of well-defined target molecules creates a bottleneck in some applications. In this paper, a carbon nanotube sensor array was constructed for the label-free discrimination of live and dead mammalian cells. Three types of carbon nanotube field-effect transistors were fabricated, and different features were extracted from the transfer characteristic curves for model training with linear discriminant analysis (LDA) and support-vector machines (SVM). Live and dead cells were accurately classified in more than 90% of samples in each sensor group using LDA as the algorithm. The recursive feature elimination with cross-validation (RFECV) method was applied to handle the overfitting and optimize the model, and cells could be successfully classified with as few as four features and a higher validation accuracy (up to 97.9%) after model optimization. The RFECV method also revealed the crucial features in the classification, indicating the participation of different sensing mechanisms in the classification. Finally, the optimized LDA model was applied for the prediction of unknown samples with an accuracy of 87.5-93.8%, indicating that live and dead cell samples could be well-recognized with the constructed model.

Pilot Verification of a Novel Approach to Remove Electrophoretic Interference of the Therapeutic Monoclonal Antibody Daratumumab.

INTRODUCTION: The advent of therapeutic monoclonal antibodies (tmAbs) in treatment of multiple myeloma poses unique challenges for the clinical laboratory. These tmAbs may appear as a detectable monoclonal protein by electrophoretic methods resulting in misinterpretation or inability to measure therapeutic responses in some patients, and there are currently limited techniques for identifying interference. In this study we performed a preliminary assessment of the SPIFE anti-daratumumab (SPIFE anti-Dara) reagent to determine whether it would be a feasible aid in resolving the interference of tmAbs with serum protein electrophoresis (SPE) and immunofixation electrophoresis (IFE). METHODS: We performed a pilot study with 20 serum samples and clinical correlates. All samples had a characteristic daratumumab electrophoretic pattern (cathodal IgG/κ). A pre-electrophoretic sample treatment was performed with SPIFE anti-Dara. The reagent is a derivatized anti-Dara that forms multiple antibody/daratumumab complexes. SPE and IFE technical procedures were performed on Helena SPIFE 3000 according to the manufacturer instructions. RESULTS: Of the 20 patients, 14 patients were identified to be on daratumumab therapy. In 14/14 of cases, the daratumumab interference was successfully removed both from SPE and IFE assays. Disease associated M-protein was still visible after pretreatment, and quantification of M-protein may be possible with the use of SPIFE anti-Dara procedure. DISCUSSION: SPIFE anti-Dara is a promising method to remove the interference of therapeutic monoclonal antibody daratumumab with SPE and IFE results in clinical laboratories and warrants further assessment.

Cerebrospinal Fluid Leak Detection with a Carbon Nanotube-Based Field-Effect Transistor Biosensing Platform.

Cerebrospinal fluid (CSF) leakage may lead to life-threatening complications if not detected promptly. However, gel electrophoresis, the gold-standard test for confirming CSF leakage by detecting beta2-transferrin (ß2-Tf), requires 3-6 h and is labor-intensive. We developed a new ß2-Tf detection platform for rapid identification of CSF leakage. The three-step design, which includes two steps of affinity chromatography and a rapid sensing step using a semiconductor-enriched single-walled carbon nanotube field-effect transistor (FET) sensor, circumvented the lack of selectivity that antitransferrin antibody exhibits for transferrin isoforms and markedly shortened the detection time. Furthermore, three different sensing configurations for the FET sensor were investigated for obtaining the optimal ß2-Tf sensing results. Finally, body fluid (CSF and serum) tests employing our three-step strategy demonstrated high sensitivity, suggesting its potential to be used as a rapid diagnostic tool for CSF leakage.

Nitrogen-Doped Carbon Nanotube Cups for Cancer Therapy.

Carbon nanomaterials have attracted significant attention for a variety of biomedical applications including sensing and detection, photothermal therapy, and delivery of therapeutic cargo. The ease of chemical functionalization, tunable length scales and morphologies, and ability to undergo complete enzymatic degradation make carbon nanomaterials an ideal drug delivery system. Much work has been done to synthesize carbon nanomaterials ranging from carbon dots, graphene, and carbon nanotubes to carbon nanocapsules, specifically carbon nanohorns or nitrogen-doped carbon nanocups. Here, we analyze specific properties of nitrogen-doped carbon nanotube cups which have been designed and utilized as drug delivery systems with the focus on the loading of these nanocapsules with specific therapeutic cargo and the targeted delivery for cancer therapy. We also summarize our targeted synthesis of gold nanoparticles on the open edge of nitrogen-doped carbon nanotube cups to create loaded and sealed nanocarriers for the delivery of chemotherapeutic agents to myeloid regulatory cells responsible for the immunosuppressive properties of the tumor microenvironment and thus tumor immune escape.

Multiplex assessment of SARS-CoV-2 antibodies improves assay sensitivity and correlation with neutralizing antibodies.

OBJECTIVES: Detection of antibodies to multiple SARS-CoV-2 antigens in a single assay could increase diagnostic accuracy, differentiate vaccination from natural disease, and aid in retrospective exposure determination. Correlation of binding antibody assessment in clinical assays with neutralizing antibodies is needed to better understand the humoral response to SARS-CoV-2 infection and establish of correlates of protection. METHODS: A cohort of 752 samples was used to assess specificity, sensitivity, and comparison to 6 other Conformitè Europëenne serologic assays for the BioRad SARS-CoV-2 IgG multiplex assay which measures receptor binding domain IgG (RBD), spike-S1 IgG (S1), spike-S2 IgG (S2), and nucleocapsid IgG (N). A subset of serial specimens from 14 patients was also tested for neutralizing antibodies (n = 61). RESULTS: Specificity for RBD and S1 IgG was 99.4% (n = 170) and 100% for S2 and N IgG (n = 170) in a cohort selected for probable interference. Overall assay concordance with other assays was >93% for IgG and total antibody assays and reached 100% sensitivity for clinical concordance at >14 days as a multiplex assay. RBD and S1 binding antibody positivity demonstrated 79-95% agreement with the presence of neutralizing antibodies. CONCLUSIONS: The BioRad SARS-CoV-2 IgG assay is comparable to existing assays, and achieved 100% sensitivity when all markers were included. The ability to measure antibodies against spike and nucleocapsid proteins simultaneously may be advantageous for complex clinical presentations, epidemiologic research, and in decisions regarding infection prevention strategies. Additional independent validations are needed to further determine binding antibody and neutralizing antibody correlations.

Differential Antibody Response to mRNA COVID-19 Vaccines in Healthy Subjects.

Knowledge about development and duration of virus-specific antibodies after COVID-19 vaccination is important for understanding how to limit the pandemic via vaccination in different populations and societies. However, the clinical utility of postvaccination testing of antibody response and selection of targeted SARS-CoV-2 antigen(s) has not been established. The results of such testing from clinical teams independent from vaccine manufacturers are also limited. Here, we report the initial results of an ongoing clinical study on evaluation of antibody response to four different SARS-CoV-2 antigens after first and second dose of Pfizer and Moderna mRNA vaccines and at later time points. We revealed a peak of antibody induction after the vaccine boosting dose with a gradual decline of antibody levels at later time. Anti-nucleocapsid antibody was not induced by spike protein-encoding vaccines and this may continue to serve as a marker of previous SARS-CoV-2 infection. No differences between the two vaccines in terms of antibody response were revealed. Age and gender dependencies were determined to be minimal within the healthy adult (but not aged) population. Our results suggest that postvaccination testing of antibody response is an important and feasible tool for following people after vaccination and selecting individuals who might require a third dose of vaccine at an earlier time point or persons who may not need a second dose due to previous SARS-CoV-2 infection. IMPORTANCE Now that authorized vaccines for COVID-19 have been widely used, it is important to understand how they induce antivirus antibodies, which antigens are targeted, how long antibodies circulate, and how personal health conditions and age may affect this humoral immunity. Here, we report induction and time course of multiple anti-SARS-CoV-2 antibody responses in healthy individuals immunized with Pfizer and Moderna mRNA vaccines. We also determined the age and gender dependence of the antibody response and compared antibody levels to responses seen in those who have recovered from COVID-19. Our results suggest the importance of screening for antibody response to multiple antigens after vaccination in order to reveal individuals who require early and late additional boosting and those who may not need second dose due to prior SARS-CoV-2 infection.