Immunoglobulin Type Research Articles

Diagnosing Autoimmune Bullous Diseases—An Indian Perspective

Abstract Introduction: Autoimmune bullous diseases (AIBDs) are a group of illnesses characterized by autoantibodies targeting adhesion molecules in the skin and mucosa. Accurate diagnosis of the specific subtype of AIBD is crucial for effective management and predicting prognosis, especially in cases with an increased risk of malignancy. However, differentiating between subtypes can be challenging due to overlapping symptoms. Overview of diagnostic tests: Direct immunofluorescence microscopy (DIF) detects in vivo bound antibodies in perilesional tissue biopsies and provides details about the probable site of autoantibody deposition within the skin/mucosae, immunoglobulin type, and pattern of antibody deposition. Indirect immunofluorescence (IIF) microscopy with organ substrate is a minimally invasive serological test that detects circulating autoantibodies. Enzyme-linked immunosorbent assay (ELISA) quantifies serum autoantibodies against specific autoantigens. Quantitative ELISA is useful for diagnosis, monitoring therapy, and assessing disease activity. Commercially available ELISA kits, including the multi-variant ones, can detect antibodies associated with AIBDs. BIOCHIP is a technique based on IIF that offers a sensitive and specific diagnostic alternative to ELISA. It uses microarrays with multiple antigenic substrates to simultaneously screen common AIBDs. The BIOCHIP slides contain different substrates, allowing the identification of multiple types of autoantibodies in a single test. Indian context: While these diagnostic tests offer valuable insights into target antigens, antibody patterns, and disease subtypes, it is important to note that the availability of these tests is limited in most centers across India. This limitation can be attributed to factors such as the relatively higher cost of these investigations, challenges related to the stability of immuno-reactants, and a shortage of trained personnel capable of performing such tests. Conclusion: This review discusses the diagnosis of AIBDs based on resources available in India, as of today. It also provides with practically applicable diagnostic algorithms for pragmatic diagnosis of AIBDs in Indian scenario.

The identification and correction of pseudohypercalcemia.

We found that a subset of patients with monoclonal gammopathy exhibited hypercalcemia without apparent causes or clinical manifestations In a cohort of 289 monoclonal gammopathy patients, 2.4% presented with such atypical hypercalcemia, with one notable case exhibiting normal ionized calcium levels, suggesting the presence of pseudohypercalcemia. The aim of this study is to elucidate the factors contributing to pseudohypercalcemia in monoclonal gammopathy and to develop a novel globulin-corrected calcium formula for clinical application. This observational study enrolled 110 monoclonal gammopathy patients from our center. An additional 33 patients were recruited to validate the newly proposed formula. Univariate analysis identified potential risk factors. And multivariate logistic regression identified definitive influential factors. The determined influential factors were utilized to develop a formula by multiple linear regression, which was validated by a paired t-test and the Youden index. This study found that globulin was a risk factor for pseudohypercalcemia. It revealed that pseudohypercalcemia should be considered in patients with globulin levels ≥61 g/L (P=0.014). Both albumin and globulin were confirmed as independent factors associated with bound calcium. Given that, we developed a formula to correct ionized calcium levels, which was consistent with ionized calcium tested by blood gas analysis. The diagnostic accuracy of the new formula (Youden index is 0.906) is better than the traditional formula (Youden index is 0.906). Interestingly, all monoclonal immunoglobulin types, except for light chains, showed an equal propensity to develop pseudohypercalcemia (P=0.306). It also showed a linear correlation between IgA, IgG, and IgM and bound calcium. This study confirmed that elevated globulin affects serum total calcium and offered the threshold of globulin ≥ 61 g/L in the differential diagnosis of peudohypercalcemia from hypercalcemia. The new formula based on albumin and globulin was developed, which was verified to be better than the traditional formula for correctly diagnosing hypercalcemia. In addition, we found that neither light chains nor heavy chains of monoclonal immunoglobulin alone can result in pseudohypercalcemia.

B-336 Analytical verification of the EXENT® solution for the identification an quantitation of monoclonal immunoglobulins by mass spectrometry

Abstract Background Recently a new mass spectrometry (MS) technology, EXENT® solution (The Binding Site, part of Thermo Fisher Scientific), for the identification and quantification of IgG, IgA and IgM intact monoclonal immunoglobulins (M-Ig) has been launched in the European markets. In this study we present the results of onsite verification study performed in our hospital. Methods Study design, using patients serum samples, including data analysis were based on the Clinical & Laboratory Standards Institute (CLSI). Relevant standards used were: Precision-CLSI EP15-A3:2014, Linearity-EP06 ED2:2020 and Comparison - EP09C ED3:2018. Samples were automatically prepared on the EXENT-iP®500 liquid handler, sample spots were analysed using the EXENT-iX®500 MALDI-TOF mass spectrometer. Light chain spectral peaks were identified by EXENT-iQ® software and then quantified indirectly in combination with total immunoglobulins concentrations by immunoturbidimetry in the Optilite® platform (The Binding Site, part of Thermo Fisher Scientific). We compared the results of MS with capillary electrophoresis (CZE) (IgG: n=82; IgA n=19; IgM n=19) and immunofixation (sIFE) (n=541). Results Precision was, for IgG with concentrations between 50.77 g/L and 3.51 g/L of 4.14% and 4.47%, for IgA between 61.88g/L and 5.02 g/L of 1.42% and 7.17%, and for IgM between 68.32 g/L and 7.30 g/L of 5.32% and 8.44%, respectively. Deviation from linearity of this assay, using serially diluted serum samples over the following ranges for IgG (65.18 g/L - 0.23 g/L), IgA (48.91 g/L - 0.26 g/L) and IgM (49.65 g/L -0.23 g/L) was <10% for all isotypes. M-Ig concentration comparison for specimens with detectable IgG, IgA or IgM M-Ig with CZE, assessed by Passing Bablok regression was IgG g/L MS = -0.9756 + 1.523x CZE g/L (slope -2.058 to -0.3322 and intercept 1.331 to 1.679), IgA g/L MS = -2.114 + 1.646x CZE g/L (slope -12.50 to -0.5059 and intercept 1.105 to 3.296), IgM g/L MS = -1.886 + 1.935x CZE g/L (slope -5.205 to -0.3571 and intercept 1.239 to 2.462). All samples MS g/L = -1.182 + 1.556x CZE g/L (n=120). Agreement between EXENT/IFE was of 99.1% for IgGK(209/211), 97.2% IgGL(105/108), 100% AK(97/97), 100% AL(51/51), 100% MK(6/6), 100% ML(3/3), 89.2% free K(33/37), 89.3% free L(25/28), 66,7% non-secretory(4/6). Discordances intact immunoglobulin was due to samples with two M-Ig, where IFE wasn’t able to identify the major clone, for free K, free L was due to EXENT’s ability to identify the heavy chain that matched with the light chain, and for non-secretory patients due to EXENT was able to identify one M-Ig in 2/6 samples. Conclusions These on-site verification results demonstrate that the EXENT solution has a wide measuring interval, to detect M proteins with very low concentrations and to provide stable and reproducible performance with a high level of accuracy for the detection and typing of monoclonal immunoglobulins. However, the bias observed against CZE to quantify M-Ig would be justified for the lowest CZE sensitivity, CZE variability and also due to the CZE quantification uses the total protein concentration to quantify M-Ig.

Smoltification of Atlantic Salmon (Salmo salar L.) Is Associated with Enhanced Traffic and Renewal of B Cell Repertoire.

The smoltification of farmed Atlantic salmon is commonly associated with mild immunosuppression. However, B cells may deviate from this trend, showing increased proliferation and migration during this period. This study assessed the effects of smoltification and adaptation to seawater in a controlled experiment. Analyses were conducted on the head kidney, spleen, gill, and both visceral and subcutaneous fat (VAT, SAT) across four time points: parr, early and complete smoltification, and twelve weeks post-seawater transfer. Gene expression analysis was performed to track the distribution and developmental changes in their B cells. Expression profiles of three types of immunoglobulins (ig), including membrane-bound and secreted forms of igm, as well as B cell-specific markers pax1 and cd79, showed strong correlations and contrasted with profiles of other immune cell markers. The highest levels of expression were observed in the lymphatic tissue, followed by the VAT. Enhanced expression in the gill and adipose tissues of smolts suggested an increase in B cell populations. Parallel sequencing of the variable region of the IgM heavy chain was used to track B cell traffic, assessed by the co-occurrence of the most abundant sequences (clonotypes) across different tissues. Smoltification markedly enhanced traffic between all tissues, which returned to initial levels after twelve weeks in the sea. The preferred migration between the head kidney, spleen, and VAT supports the role of abdominal fat as a reservoir of lymphocytes. These findings are discussed in the context of recent studies that suggested the functional significance of B cell traffic in Atlantic salmon. Specifically, the migration of B cells expressing secreted immunoglobulins to virus-infected hearts has been identified as a key factor in the disease recovery and survival of fish challenged with salmon alphavirus (SAV); this process is accelerated by vaccination. Additionally, the study of melanized foci in the skeletal muscles revealed an association between antigen-dependent differentiation and the migration of B cells, indicating a transfer from local to systemic immune responses. Updating the antibody repertoire in the lymphatic and peripheral tissues of smolts may assist in their adaptation to the marine environment and in encountering new pathogens. Emerging evidence highlights B cell migration as an important and previously unrecognized immune mechanism in salmonids.

Transamniotic Fetal Immunotherapy with Secretory IgA: A Potential Novel Ancillary Strategy for the Prevention of Necrotizing Enterocolitis

Plain Language SummaryThis is an experimental study in an animal model showing that a special type of immunoglobulin called secretory immunoglobulin-A (SIgA), which can find its way to the fetal intestine after a simple injection into the amniotic fluid, is capable of attaching itself to intestinal bacteria after birth. The importance of that is the fact that SIgA has been previously shown to protect neonates, especially premature ones, from a dangerous disease called necrotizing enterocolitis (NEC). Since certain premature neonates cannot take SIgA by mouth, this new form of delivery of that immunoglobulin could become a novel minimally invasive alternative to give babies that molecule and hopefully help protect them from NEC. Further studies need to be performed before this can be used in humans, yet the original data presented in this study justify said studies while also suggesting that transamniotic fetal immunotherapy with SIgA could become a novel strategy for prevention of not only NEC but also possibly other perinatal infections.

Correlation between Morphological Typing and Monoclonality of Bone Marrow Plasma Cells and Its Diagnostic Value for High-Risk Smoldering Multiple Myeloma

To investigate the correlation between morphological typing and monoclonality of bone marrow plasma cells, and explore the diagnostic value of plasma cell morphological typing for high-risk smoldering multiple myeloma(HR-SMM). The correlation between the morphological characteristics and the monoclonality of bone marrow plasma cells was analyzed in 84 patients with HR-SMM who treated in our hospital. The consistency of morphologically abnormal bone marrow plasma cells with serum free light chain (sFLC) ratio, next-generation sequencing (NGS) detection results, and its correlation with monoclonal plasma cells detected by flow cytometry (FCM) were further verified. The immunoglobulin types and levels of non-involved immunoglobulins in serum of the patients were detected, and the distribution of plasma cell clusters in patients with different disease was observed. The mean percentage of mature plasma cells were decreased successively in the order of reactive plasmacytosis (RP) group, monoclonal gammopathy of undetermined significance (MGUS) group, smoldering multiple myeloma (SMM) group, HR-SMM group and multiple myeloma (MM) group; while the mean percentage of immature, primitive, reticular and flaming plasma cells were increased successively in the order of RP group, MGUS group, SMM group, and HR-SMM group, and the difference between any two groups was statistically significant (P < 0.05).The average proportion of abnormal plasma cells in the bone marrow of HR-SMM patients was 96.2% of the total plasma cells. The proportion of abnormal plasma cells were in good agreement with the sFLC ratio and the results of NGS detection in HR-SMM patients (kappa=0.879 and kappa=0.891, both >0.75)，and showed good correlation with the monoclonal plasma cells with immunophenotype of CD45-/CD38+/CD138+/CD56+/CD19-( γ=0.825). The levels of non-involved immunoglobulin in IgG, IgA and IgM type HR-SMM patients were all decreased by more than 25% compared with the normal reference range, and the differences were statistically significant (P < 0.05). There was no significant difference in the distribution ratio of plasma cell clusters among different disease groups (P >0.05). In HR-SMM patients, the immature, primitive, reticular and flaming plasma cells in bone marrow are considered as abnormal plasma cells, and they are correlated with monoclonal plasma cells. The proportion of abnormal plasma cells in total plasma cells of bone marrow and the reduction extent of non-involved immunoglobulin level in patients have certain reference value for the diagnosis of HR-SMM.

Determinants of Systemic SARS-CoV-2-Specific Antibody Responses to Infection and to Vaccination: A Secondary Analysis of Randomised Controlled Trial Data.

SARS-CoV-2 infections elicit antibodies against the viral spike (S) and nucleocapsid (N) proteins; COVID-19 vaccines against the S-protein only. The BCG-Corona trial, initiated in March 2020 in SARS-CoV-2-naïve Dutch healthcare workers, captured several epidemic peaks and the introduction of COVID-19 vaccines during the one-year follow-up. We assessed determinants of systemic anti-S1 and anti-N immunoglobulin type G (IgG) responses using trial data. Participants were randomised to BCG or placebo vaccination, reported daily symptoms, SARS-CoV-2 test results, and COVID-19 vaccinations, and donated blood for SARS-CoV-2 serology at two time points. In the 970 participants, anti-S1 geometric mean antibody concentrations (GMCs) were much higher than anti-N GMCs. Anti-S1 GMCs significantly increased with increasing number of immune events (SARS-CoV-2 infection or COVID-19 vaccination): 104.7 international units (IU)/mL, 955.0 IU/mL, and 2290.9 IU/mL for one, two, and three immune events, respectively (p < 0.001). In adjusted multivariable linear regression models, anti-S1 and anti-N log10 concentrations were significantly associated with infection severity, and anti-S1 log10 concentration with COVID-19 vaccine type/dose. In univariable models, anti-N log10 concentration was also significantly associated with acute infection duration, and severity and duration of individual symptoms. Antibody concentrations were not associated with long COVID or long-term loss of smell/taste.

Highly-Sensitive Polymer Optical Fiber SPR Sensor for Fast Immunoassay

A new type of human immunoglobulin G (IgG) sensors based on the surface plasmon resonance (SPR) in the low refractive index (RI) plastic optical fiber (POF) and an antibody immobilization method is presented. A 50-nm-thick gold film was formed on the polished D-shaped fiber surface by magnetron sputtering. The RI response of the POF sensor is 30 049.61 nm/RIU, which is 26.5 times higher than that of single mode fiber (SMF) SPR sensors. The proposed SPR biosensor can be developed by simple and rapid modification of the gold film with 11-mercapto undecanoic acid (MUA). Upon immobilization of the goat anti-human IgG antibody, the resonance wavelength shifts by 11.2 nm. The sensor can be used to specifically detect and quantify the human IgG at concentrations down to 245.4 ng/mL with the sensitivity of 1.327 7 nm per µg/mL, which offers an enhancement of 12.5-fold compared to that of the conventional SMF based SPR sensors. The proposed device may find the potential applications in the case of use at the point of care.

Antibody-displaying extracellular vesicles for targeted cancer therapy.

Extracellular vesicles (EVs) function as natural delivery vectors and mediators of biological signals across tissues. Here, by leveraging these functionalities, we show that EVs decorated with an antibody-binding moiety specific for the fragment crystallizable (Fc) domain can be used as a modular delivery system for targeted cancer therapy. The Fc-EVs can be decorated with different types of immunoglobulin G antibody and thus be targeted to virtually any tissue of interest. Following optimization of the engineered EVs by screening Fc-binding and EV-sorting moieties, we show the targeting of EVs to cancer cells displaying the human epidermal receptor 2 or the programmed-death ligand 1, as well as lower tumour burden and extended survival of mice with subcutaneous melanoma tumours when systemically injected with EVs displaying an antibody for the programmed-death ligand 1 and loaded with the chemotherapeutic doxorubicin. EVs with Fc-binding domains may be adapted to display other Fc-fused proteins, bispecific antibodies and antibody-drug conjugates.

Presence of immunoglobulin E-expressing antibody-secreting cells in the dermis close to bullous pemphigoid lesions.

Antibody-secreting cells (ASCs) produce immunoglobulin (Ig) G and IgE autoantibodies in secondary lymphoid organs. Evidence also suggests their existence in the skin in various chronic inflammatory conditions, and in association with CXCL12 and CXCL13, they regulate the recruitment/survival of ASCs and germinal center formation to generate ASCs, respectively. However, the presence of IgG and IgE in bullous pemphigoid (BP) lesions needs to be addressed. Here, we aimed to analyse BP skin for the presence of IgG and IgE and the factors contributing to their generation, recruitment, and persistence. Skin samples from 30 patients with BP were stained to identify ASCs and the immunoglobulin type they expressed. The presence of tertiary lymphoid organ (TLO) elements, which generate ASCs in non-lymphoid tissues, and the chemokines CXCL12 and CXCL13, which regulate the migration/persistence of ASCs in lymphoid tissues and formation of TLOs, respectively, were evaluated in BP skin. BP skin harboured ASCs expressing the two types of antibodies IgG and IgE. ASCs were found in high-grade cellular aggregates containing TLO elements: T cells, B cells, CXCL12+ cells, CXCL13+ cells and high endothelial venules. IgG+ ASCs were detected among these aggregates, whereas IgE+ ASCs were dispersed throughout the dermis. CXCL12+ fibroblast-like cells were located close to ASCs. The inflammatory microenvironment of BP lesions may contribute to the antibody load characteristic of the skin of patients with BP by providing a site for the presence of ASCs. CXCL13 and CXCL12 expression may contribute to the generation and recruitment/survival of ASCs, respectively.

Electric Field-Assisted Molecularly Imprinted Polymer-Modified QCM Sensor for Enhanced Detection of Immunoglobulin.

In this study, an electric-field-assisted molecularly imprinted polymer (EFAMIP) as an enhanced form of MIP was developed to improve the MIP-modified quartz crystal microbalance (QCM) biosensors. While exerting a vertical electric field, polymerization of methacrylic acid in the presence of immunoglobulin G (IgG) as the template was initiated, and later, after the template removal process, the EFAMIPs were obtained. The polymer surface characterization was conducted by using a scanning electron microscope. The impact of electric field direction on IgG binding sites, forming either EFAMIP-Fab or EFAMIP-Fc, was assessed. Next, the static measurement results in liquid for EFAMIP-modified QCM and MIP-modified QCM were compared. While encompassing IgG, EFAMIP-modified QCMs exhibited up to a 113.5% higher frequency shift than typical MIP in time-limited detection. The final frequency shift of EFAMIP, which determines the detection limit of IgG, was improved up to 12.5% compared to typical MIP. Moreover, the EFAMIP-Fab performance was promising for the selective detection of IgG in a solution containing different types of immunoglobulins.

The Associations between Exposure to Multiple Heavy Metals and Total Immunoglobulin E in U.S. Adults.

Immunoglobulin E (IgE) is a type of immunoglobulin, and elevated serum total IgE is often present in allergic diseases. Exposure to environmental heavy metals has been markedly linked to allergic diseases, leading to elevated total IgE levels. However, studies concerning the effects of multiple metal exposures on total IgE levels are limited. Therefore, the current study seeks to explore the correlation between heavy-metal co-exposure and total IgE levels based on the National Health and Nutrition Examination Survey (NHANES, 2005-2006). Participants possessed complete data on total IgE levels, 11 urinary metal concentrations and other covariates. The correlations between 11 metals and total IgE levels were analyzed using multiple linear regression, and total IgE levels were a continuous variable. Total IgE levels exceeding 150 kU/L were considered sensitized. Binary logistic regression analyses were employed to assess the correlation between metal exposure and the occurrence of an allergic state. Then, the association between co-exposure to the 11 metals and total IgE levels or the occurrence of sensitization status was further analyzed by Bayesian kernel machine regression (BKMR), a multi-contaminant model. There were 1429 adults with complete data included. Based on the median concentration, molybdenum (Mo) had the highest concentration (46.60 μg/L), followed by cesium (Cs), barium (Ba), lead (Pb), and mercury (Hg). And the median (interquartile range) for total IgE levels was 43.7 (17.3, 126.0) kU/L. Multiple linear regression results showed that Pb was significantly and positively associated with total IgE levels (β = 0.165; 95% CI: 0.046, 0.284). Binary logistic regression showed a significant positive correlation between urinary Pb (OR: 1.258; 95% CI: 1.052, 1.510) and tungsten (W) (OR: 1.251; 95% CI: 1.082, 1.447). Importantly, the BKMR model found a positive correlation between combined-metal exposure and total IgE levels and the occurrence of sensitization status. The mixed heavy-metal exposure was associated with increased total IgE levels, and this association may be driven primarily by the exposure of Pb and W. This study provides new insights into the relationship between heavy-metal exposure and allergic diseases. More research is needed to confirm these findings.

Immunity transfer in mule foals fed with good IgG quality colostrum

While the passive transfer of immunity in horse and donkey foals has been extensively studied, there is limited information for mule foals. Immunoglobulin type G (IgG) and serum total protein concentration (TP) were assessed at different sampling times to evaluate the correlation between serum radial immunodiffusion (SRID) with electrophoresis, refractometry, and dry chemistry analyzer (Biuret), and to estimate serum IgG concentrations using serum TP in mule foals. We analyzed a total of 30 samples collected at birth, and at 6, 12, 24, and 48 h of life from 6 mule foals by SRID, electrophoresis TP, biuret TP, and refractometry TP.The SRID IgG concentration significantly increased from birth until T6 (p < 0.001). Serum TP analyzed with refractometry revealed differences between T0 and T12, T24 and T48 (p < 0.05), while a significant difference was observed with the biuret method between T0 and all the other sampling times (p < 0.001).A strong correlation was found between IgG SRID and biuret TP (r = 0.69, p < 0.001), and a good correlation existed between IgG SRID, refractometry TP, and electrophoresis TP (r = 0.44, p < 0.01 and r = 0.39, p < 0.05, respectively).All methods can be used to estimate the passive transfer of immunity in mule foals. TP refractometry and biuret TP values can be used to determine serum IgG concentrations in the blood of mule foals on their first day of life through the application of a specific equation.

Anti-SARS-CoV-2-specific antibodies in human breast milk following SARS-CoV-2 infection during pregnancy: a prospective cohort study

BackgroundWhile the presence of SARS-CoV-2 in human breast milk is contentious, anti-SARS-CoV-2 antibodies have been consistently detected in human breast milk. However, it is uncertain when and how long the antibodies are present.MethodsThis was a prospective cohort study including all consecutive pregnant women with confirmed SARS-CoV-2 infection during pregnancy, recruited at six maternity units in Spain and Hong Kong from March 2020 to March 2021. Colostrum (day of birth until day 4 postpartum) and mature milk (day 7 postpartum until 6 weeks postpartum) were prospectively collected, and paired maternal blood samples were also collected. Colostrum samples were tested with rRT-PCR-SARS-CoV-2, and skimmed acellular milk and maternal sera were tested against SARS-CoV-2 specific immunoglobulin M, A, and G reactive to receptor binding domain of SARS-CoV-2 spike protein 1 to determine the presence of immunoglobulins. Then, we examined how each immunoglobulin type in the colostrum was related to the time of infection by logistic regression analysis, the concordance between these immunoglobulins in the colostrum, maternal serum, and mature milk by Cohen's kappa statistic, and the relationship between immunoglobulin levels in mature milk and colostrum with McNemar.ResultsOne hundred eighty-seven pregnant women with confirmed SARS-CoV-2 infection during pregnancy or childbirth were recruited and donated the milk and blood samples. No SARS-CoV-2 was found in the human breast milk. Immunoglobulin A, G, and M were present in 129/162 (79·6%), 5/163 (3·1%), and 15/76 (19·7%) colostrum samples and in 17/62 (27·42%), 2/62 (3·23%) and 2/62 (3·23%) mature milk samples, respectively. Immunoglobulin A was the predominant immunoglobulin found in breast milk, and its levels were significantly higher in the colostrum than in the mature milk (p-value < 0.001). We did not find that the presence of immunoglobulins in the colostrum was associated with their presence in maternal, the severity of the disease, or the time when the infection had occurred.ConclusionsSince anti-SARS-CoV-2 antibodies are found in the colostrum irrespective of the time of infection during pregnancy, but the virus itself is not detected in human breast milk, our study found no indications to withhold breastfeeding, taking contact precautions when there is active disease.

Neuroimmune communication in allergic rhinitis.

The prevalence rate of allergic rhinitis (AR) is high worldwide. The inhalation of allergens induces AR, which is an immunoglobulin E-mediated and type 2 inflammation-driven disease. Recently, the role of neuroimmune communication in AR pathogenesis has piqued the interest of the scientific community. Various neuropeptides, such as substance P (SP), vasoactive intestinal peptide (VIP), calcitonin gene-related peptide (CGRP), nerve growth factor (NGF), and neuromedin U (NMU), released via "axon reflexes" or "central sensitization" exert regulatory effects on immune cells to elicit "neurogenic inflammation," which contributes to nasal hyperresponsiveness (NHR) in AR. Additionally, neuropeptides can be produced in immune cells. The frequent colocalization of immune and neuronal cells at certain anatomical regions promotes the establishment of neuroimmune cell units, such as nerve-mast cells, nerve-type 2 innate lymphoid cells (ILC2s), nerve-eosinophils and nerve-basophils units. Receptors expressed both on immune cells and neurons, such as TRPV1, TRPA1, and Mas-related G protein-coupled receptor X2 (MRGPRX2) mediate AR pathogenesis. This review focused on elucidating the mechanisms underlying neuroimmune communication in AR.

Features of the functioning of the natural defense mechanisms of piglets under the influence of immunotropic substances

The number of studies related to detailed characteristics of immune system development in early stages of ontogenesis are directed to expansion of concepts of mechanisms of inherent and adaptive immunity in premature newborns. However, discrepancy in publication data and absence of clear conceptions about unique features of immune system in newborn piglets for different gestation periods have significant actuality for further investigation. The study was carried out to establish characteristics of phenotypic and functional characteristics of the immune cells and establishment of adaptive immunity in piglets. The antibody repertoire of sow has interested immunologists for decades, in part because of the ease with which large quantities of high affinity antibodies can be observed in serum, and into other fluids because of the presence of genetic variants for both light and heavy chain of all known immunoglobulin types. The integrative analysis of the cellular and humoral immunity in piglets during early ontogenesis carried out to clarify the exact mechanisms of the immune response modulation. Effectiveness and selectivity of the immunotropic chemicals application to regulate immune cell populations in piglets is presented. The progress in immune cell populations is regulated by the various cytokines production in the universal concentration-dependent manner. Furthermore, the development of the immune functions cross reacts with innate immunity mechanisms including anti-bacterial enzymes, epithelial barrier integrity in the both skin and intestine. The interaction between multiple components of innate immunity is a critic initiator of the dynamic changes in adaptive immunity. The natural resistance targets to the establishment of infection root and delays disease progression. The duration of the exposure to colostrum in important role in the dynamics of immune response as well as its efficacy. The chapter discusses the plural mechanisms so far proposed to be responsible for the modulation natural resistance.

Adhesion of Immunoglobulins to Band3 Promotes Increased Erythrocyte Sedimentation Rate in Multiple Myeloma

An increase in ESR can be seen in approximately 85% of patients with MM [1]. Therefore, ESR is often used as one of the sensitive indicators of active MM [1]. It is now widely accepted that the increase in ESR is related to the adhesion of immunoglobulins secreted by myeloma cells to erythrocytes [2, 3]. However, this explanation is descriptive and speculative and has not been confirmed by experimental data [3]. We applied peripheral blood erythrocyte samples from MM patients and used immunofluorescence/immunoprecipitation tandem protein analysis/molecular docking to determine the molecular mechanism by which all types of MM immunoglobulins can target and adhere to erythrocyte membrane Band3. This result, not only helps to explain the molecular mechanism of ESR acceleration in MM, but will also advance our understanding of the pathophysiologic changes that occur in ESR. At the same time, it may provide a new way to target protection of normal erythrocytes and targeted removal of erythrocyte-bound immunoglobulins.

B-181 Multi-centre Analytical Performance Evaluation of the EXENTâ Solution for Identification and Quantitation of Monoclonal Immunoglobulins

Abstract Background Current routine methodologies for monoclonal immunoglobulin measurements may not be sufficiently sensitive to reflect the depth of response seen in patients since the introduction of novel therapies; mass spectrometry may offer a valuable, sensitive alternative approach. Here we describe the preliminary analytical performance characteristics of the EXENT® solution (in development by The Binding Site, part of Thermo Fisher scientific) that combines specific immunoprecipitation steps and mass spectrometry for the identification and quantification of IgG, IgA and IgM intact monoclonal immunoglobulins. Each intact monoclonal immunoglobulin clone can be tracked using its unique m/z value. Methods The Lower Limit of Measuring Interval (LLMI) was established for each immunoglobulin type following EP17-A2:2012. Linearity studies were performed according to CLSI EP06-A2:2020 using high and low pools of IgG, IgA, and IgM monoclonal samples and with additional linearity testing below 1 g/L for each specificity to more effectively demonstrate low-end linearity. Within run, between run, between analyzer, between lot and total precision for M protein concentrations and for molecular mass (m/z) of the monoclonal peaks were assessed according to CLSI EP5-A3-2014. Interference was tested following ED3:2018 using 20 potential interferents against 7 samples including high and low IgG, IgA and IgM monoclonal samples. Results The initial indication of the EXENT solution performance, in development, are set out below. Results suggest an analytical sensitivity of the assays around 15 mg/L at the LLMI for all specificities. Linearity over a range of 0.014–88.9 g/L for IgG, 0.011–68.4 g/L for IgA, and 0.11–74.2 g/L for IgM. Coefficients of variation (CVs) for M protein concentrations in precision studies were &amp;lt;15% for all specificities and samples. Mass/charge values were within ±1.1 to ±1.5 m/z in total precision studies, and within ±2.7 to ±3.9 m/z in between lot precision studies, respectively, for M proteins with an m/z value ranging from 11 360.6 to 11 698.5 m/z. No significant interference effects were observed when testing the 20 interferents including intralipid (20 g/L), triglyceride (15 g/L), bilirubin (400 mg/L), rheumatoid factor (200 IU/mL) and haemoglobin (10 g/L). Conclusion The new EXENT solution demonstrates the potential for a wide measuring interval and the ability to detect M proteins with very low concentration. Also, it could provide stable and reproducible performance for the detection and typing of monoclonal immunoglobulins.

Shark IgNAR: The Next Broad Application Antibody in Clinical Diagnoses and Tumor Therapies?

Antibodies represent a relatively mature detection means and serve as therapeutic drug carriers in the clinical diagnosis and treatment of cancer-among which monoclonal antibodies (mAbs) currently occupy a dominant position. However, the emergence and development of small-molecule monodomain antibodies are inevitable due to the many limitations of mAbs, such as their large size, complex structure, and sensitivity to extreme temperature, and tumor microenvironments. Thus, since first discovered in Chondroid fish in 1995, IgNAR has become an alternative therapeutic strategy through which to replace monoclonal antibodies, thus entailing that this novel type of immunoglobulin has received wide attention with respect to clinical diagnoses and tumor therapies. The variable new antigen receptor (VNAR) of IgNAR provides an advantage for the development of new antitumor drugs due to its small size, high stability, high affinity, as well as other structural and functional characteristics. In that respect, a better understanding of the unique characteristics and therapeutic potential of IgNAR/VNAR in clinical and anti-tumor treatment is needed. This article reviews the advantages of its unique biochemical conditions and molecular structure for clinical diagnoses and novel anti-tumor drugs. At the same time, the main advantages of the existing conjugated drugs, which are based on single-domain antibodies, are introduced here, thereby providing new ideas and methods for the development of clinical diagnoses and anti-tumor therapies in the future.

Structure determination and analysis of titin A-band fibronectin type III domains provides insights for disease-linked variants and protein oligomerisation

Titin is the largest protein found in nature and spans half a sarcomere in vertebrate striated muscle. The protein has multiple functions, including in the organisation of the thick filament and acting as a molecular spring during the muscle contraction cycle. Missense variants in titin have been linked to both cardiac and skeletal myopathies. Titin is primarily composed of tandem repeats of immunoglobulin and fibronectin type III (Fn3) domains in a variety of repeat patterns; however, the vast majority of these domains have not had their high-resolution structure determined experimentally. Here, we present the crystal structures of seven wild type titin Fn3 domains and two harbouring rare missense variants reported in hypertrophic cardiomyopathy (HCM) patients. All domains present the typical Fn3 fold, with the domains harbouring variants reported in HCM patients retaining the wild-type conformation. The effect on domain folding and stability were assessed for five rare missense variants found in HCM patients: four caused thermal destabilization of between 7 and 13 °C and one prevented the folding of its domain. The structures also allowed us to locate the positions of residues whose mutations have been linked to congenital myopathies and rationalise how they convey their deleterious effects. We find no evidence of physiological homodimer formation, excluding one hypothesised mechanism as to how titin variants could exert pathological effects.