Detection Of Antibodies Research Articles

Determinants of protective humoral response to mRNA-1273 and BNT162b2 vaccines in peritoneal dialysis patients: a prospective cohort study

Patients with chronic kidney disease (CKD) on dialysis have a higher mortality rate associated with SARS-CoV-2 infection. Although vaccines are now available, the protective response rates and determinants of humoral response to the vaccine are poorly described in patients on peritoneal dialysis. This was a prospective observational study describing the response rates of detectable and standardized protective antibody titers one month after each mRNA vaccine dose in a cohort of 88 patients on peritoneal dialysis. We found that the vast majority of patients produced protective levels of antibodies (73%) one month after the second vaccine dose. In the multivariate analysis, the single determinant for an adequate humoral response was the weekly Kt/V, a surrogate of dialysis dose. The response rate was higher, but not significantly, with the mRNA-1273 than with the BNT162b2 vaccine one month after the second dose (78.7 vs. 46.2%, respectively, p = 0.02). We found that patients on peritoneal dialysis had a satisfactory humoral response rate, which was much higher than in transplant recipients. PD patients with a poor humoral response, particularly those with a low wKT/V, may benefit from an additional dose of vaccine.

Nanopore-assisted ELISA for ultrasensitive, portable, and on-site detection of ricin

Effective detection technologies in food safety with the merits of portable and on-site detection potential are always in pressing demand. Herein, we developed a nanopore-assisted Enzyme-linked immunosorbent assay (NELISA) platform, which innovatively introduced hairpin DNA (HP DNA) probes as reaction substrates. This innovation of substrates effectively avoided the inherent limitations of colorimetric signals (i.e., low sensitivity and inaccurate results) and greatly improved the sensitivity and accuracy of NELISA platform. The alkaline phosphatase (ALP)-modified detection antibody (ALP-Ab2) can specifically bind to ricin and hydrolyze the phosphate groups modified on the HP DNA probes. Nanopore recordings demonstrated that two states of probes produced highly distinguishable nanopore events, enabling the qualitative and quantitative detection of ricin. This NELISA platform fully combined the specificity of ELISA with the ultra-sensitivity, and unique single-molecule fingerprint recognition of nanopore, showing a great on-site detection potential. This method achieved the ultrasensitive and reliable detection of ricin down to 2.46 fg/mL, which enhanced the detection sensitivity by at least 106-fold compared to traditional ELISA. Furthermore, the proposed method was capable of accurately detecting ricin in real food samples with satisfactory recoveries.

Evaluation of Tissue Tropism and Horizontal Transmission of a Duck Enteritis Virus Vectored Vaccine in One-Day-Old Chicken

Herpesvirus of turkey (HVT) recombinant vector vaccines are widely used in the poultry industry. However, due to limitations in loading multiple foreign antigens into a single HVT vector, other viral vectors are urgently needed. Since chickens lack maternal immunity to duck enteritis virus (DEV), vector vaccines using DEV as a backbone are currently under study. Even though a recently developed DEV vector vaccine expressing the influenza hemagglutinin H5 of highly pathogenic avian influenza (DEV-H5) induces highly detectable anti-HA antibodies, safety issues hamper further vaccine development. In this work, tissue affinity and horizontal transmission in 1-day-old chickens were systematically evaluated after DEV-H5 vector vaccine inoculation. Sixty percent of DEV-H5-inoculated chickens died between day 2 and day 7 post-inoculation. The displayed clinical signs consisted of lethargy, anorexia, and diarrhea, and virus was shed in feces. Gross and/or histological lesions were recorded in the kidney, heart, intestine, liver, lung, and spleen. Moreover, DEV-H5 replication in intestinal cells caused an increment in interferon-α expression, while occluding junction proteins and ZO-1 expression were significantly upregulated. As a control, birds inoculated with a commercial recombinant turkey herpesvirus expressing the VP2 protein of the infectious bursal disease virus (HVT-VP2) vector vaccine showed neither clinical signs nor mortality. Overall, while the HVT-VP2 vaccine demonstrated complete safety in 1-day-old chickens, our potential DEV-H5 vaccine requires further attenuation for consideration as a vector vaccine candidate in chickens.

Detection of Bovine Respiratory Syncytial Virus in Cattle: A Systematic Review and Meta-Analysis

Bovine respiratory syncytial virus (BRSV) is an economically important pathogen of cattle and contributes to the bovine respiratory disease complex (BRDC). Despite individual studies investigating BRSV prevalence, risk factors, and detection methodologies, a systematic review and meta-analysis have been lacking. The aim of the current study was to conduct a systematic review and meta-analysis to determine the prevalence and detection rate of BRSV and identify associated risk factors. Additionally, the study aimed to explore the variability in BRSV prevalence based on different detection methods and associated risk factors. Adhering to PRISMA guidelines, data from three databases—Web of Science, PubMed, and Scopus—were systematically retrieved, screened and extracted. Out of 2790 initial studies, 110 met the inclusion criteria. The study found that prevalence and detection rates varied based on the detection methods used (antibody, antigen, and nucleic acid), study populations, production systems, and geographic locations. Findings were reported as a pooled proportion. The pooled proportion, hereafter referred to as prevalence or detection rate, was determined by calculating the ratio of cattle that tested positive for BRSV to the total number of cattle tested. Key findings include a pooled prevalence of 0.62 for antibody-based methods, 0.05 for antigen-based methods, and 0.09 (adjusted to 0.03) for nucleic acid-based methods. Detection rates in BRDC cases also varied, with antibody methods showing a rate of 0.34, antigen methods 0.16, and nucleic acid methods 0.13. The certainty of evidence of the meta-analysis results, assessed using GRADE, was moderate for antibody detection methods and low for antigen and nucleic acid methods. The study identified significant risk factors and trends affecting BRSV prevalence, such as geographical location, herd size, age, and co-infections. The results of the current study showed the complexity of understanding BRSV prevalence in different settings. The variability in BRSV prevalence based on detection methods and associated risk factors, such as geographic location and herd size, highlights the need for tailored approaches to detect and manage BRSV accurately.

Ultrasensitive ECL immunoassay for CA15-3 via self-enhanced L012-loaded ZnNi-MOF as an emitter and CeO2–Pt as a co-reaction accelerator

Improvement in the sensitivity and accuracy of ECL-based biosensors, to a great extent, depends on the fabrication of high-performance electrochemiluminescence (ECL) sensors. Herein, polyethyleneimine (PEI)-grafted bimetallic ZnNi metal-organic framework (ZnNi-MOF/PEI) nanospheres were synthesized to prepare a L012-based ECL immunosensor for detecting carbohydrate antigen 15-3 (CA15-3). The introduced PEI was utilized not only as a bridging element for L012 luminophore and detection antibody (Ab2) immobilization but also as a co-reactant for generating the self-enhanced ZnNi-MOF/PEI-L012@Ab2 ECL signal probe. Furthermore, Pt nanoparticle-functionalized CeO2 nanorods (CeO2–Pt) featuring high-efficiency electrocatalytic properties, as a co-reaction accelerator, could catalyze H2O2 to generate abundant reactive oxygen species (ROS), further augmenting the ECL signal of ZnNi-MOF/PEI-L012@Ab2. In the presence of CA15-3 antigen, the capture antibody (Ab1)-conjugated CeO2–Pt complex (CeO2–Pt@Ab1) and ZnNi-MOF/PEI-L012@Ab2 would get closer to each other, with the formation of a “signal-on” sandwich-type ECL immunosensor. The ZnNi-MOF-assisted ECL biosensor achieved a detection limit of down to 5.75✕10−5 U mL−1. This work shows distinguished potential for measuring low levels of biomarkers that may help in the clinical diagnosis of breast cancer.

Infant feeding and treatment practices could lead to enhanced transmission of Kaposi’s sarcoma-associated herpesvirus (KSHV) and other orally shed infections via saliva, in rural south-western Uganda

ABSTRACT This qualitative sub-study investigated household practices affecting orally shed infections using Kaposi’s sarcoma-associated herpesvirus (KSHV) as a focus. Participants enrolled from 50 households in rural south-western Uganda were followed monthly up to three times. At enrolment, in-depth interviews were completed, and venous blood collected. KSHV seropositivity was defined as anti-KSHV antibody detection to any of 25 antigens by multiplex bead-based assay. Mouthwash samples from every visit were tested by qPCR and KSHV shedders defined as individuals with KSHV DNA detected. At least one KSHV seropositive person was in 48/49(98%) households. Among those, 79% had 1+ KSHV shedders including 45% with 1+ always shedders and 92% with 1+ intermittent shedders, not mutually exclusively. All respondents reported feeding infants with pre-masticated hard food/fruits and testing food/tea temperature. Temperature was tested by tasting, pouring tea on their hand, or touching the cup to their cheek. Some cooled food/tea using a utensil or blowing over it. Food sharing amongst children and adults and using the same dish was common practice. To treat colic pain, carers/mothers reported chewing herbs and spitting into the child’s mouth. Feeding and treatment practices did not vary by KSHV status. We identified potential KSHV transmission modes in rural Ugandan households.

Expression of Immunodominant gI Protein of Duck Enteritis Virus and its Evaluation for ELISA

Background: Duck plague (DP), also known as duck viral enteritis (DVE) caused by Anatid alpha herpesvirus-1, is the major infectious viral disease reported in India very often with significant economic losses due to high morbidity and mortality. The genome is approximately 158,091 bp with a genomic arrangement pattern (UL-IRS-US-TRS) that corresponds to type-D herpesvirus. The incubation period of the disease in domestic ducks ranges from 3 to 7 days and the mortality varies from 5-100%. It has been reported that the strategies to monitor and control DVE were often frustrating because the disease tends to establish a latent infection in waterfowl and vaccination failure. The present study was taken as an attempt to develop indirect ELISA using developed recombinant gI protein to diagnose DEV in field condition. Methods: In this study, we selected a membrane glycoprotein protein I (US7) gene, that plays an important role in virion sorting, cell-cell spread and binding of IgG Fc receptor. The gI protein was cloned with pET-32a (+) and expressed in a prokaryotic (E. coli) expression system. Further, recombinant protein was purified by nickel column affinity chromatography and refolded by dialysis. The obtained concentrated protein was then evaluated for its antigenicity and reliability in an Indirect-Enzyme-linked immunosorbent assay (ELISA) for DEV antibody detection in duck sera. Result: A panel of positive, negative and vaccinal serum was evaluated which indicated the potential use of the protein in the development of a serological assay for sero- surveillance of duck plague infection.

The Impact of Reduced Immunosuppression on Alloimmunity: A Retrospective Study of Pediatric Kidney Transplant Recipients.

Kidney transplantation is the best treatment for end-stage kidney disease but requires immunosuppressive medications, which have significant side effects. Many pediatric recipients experience these side effects, leading to dosage reductions, which potentially increase the risk of alloimmunity. We aimed to describe the alteration in immunosuppressive medication, explore the reasons for the reductions, and assess the potential impact on alloimmunity. Data from 49 pediatric kidney transplant recipients receiving an allograft from 2009 to 2020 were retrospectively studied. The recipients were screened for HLA antibodies after the transplantation. The median age of recipients was 11 years (IQR 8), with a median follow-up of 5 years (IQR 5). Eighty percent of the transplantations were corticosteroid-free. During follow-up, 11% developed de novo donor-specific antibodies (dnDSA), and 60% had detectable HLA antibodies. The 1-year rejection rate was 4%. Immunosuppressive medication was altered substantially in most recipients, resulting in 72% being on mono- or dual therapy with a reduced mycophenolate mofetil (MMF) dosage by the end of the first posttransplant year. The median MMF dose was nearly half of the intended. Tacrolimus levels were maintained close to the target of 5 ng/mL. No association was found between reduced immunosuppression and dnDSA or rejections. Reductions were primarily due to MMF-related side effects: leukopenia in 77%, gastrointestinal issues in 34%, and infections with Epstein-Barr virus, cytomegalovirus, and BK polyomavirus in 49%. Reduced MMF with a sufficient trough tacrolimus level in a population of mainly corticosteroid-free pediatric kidney transplant recipients did not lead to unacceptable alloimmunity.

Repeated COVID-19 mRNA-based vaccination contributes to SARS-CoV-2 neutralizing antibody responses in the mucosa.

To prevent infection by respiratory viruses and consequently limit virus circulation, vaccines need to promote mucosal immunity. The extent to which the currently used messenger RNA (mRNA)-based COVID-19 vaccines induce mucosal immunity remains poorly characterized. We evaluated mucosal neutralizing antibody responses in a cohort of 183 individuals. Participants were sampled at several time points after primary adenovirus vector-based or mRNA-based COVID-19 vaccination and after mRNA-based booster vaccinations. Our findings revealed that repeated vaccination with mRNA boosters promoted severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) neutralizing antibodies in nasal secretions. Nasal and serum neutralizing antibody titers of both IgG and IgA isotypes correlated to one another. We investigated the source of these mucosal antibodies in a mouse model wherein mice received repeated mRNA vaccines for SARS-CoV-2. These experiments indicated that neutralizing antibody-producing cells reside in the spleen and bone marrow, whereas no proof of tissue homing to the respiratory mucosa was observed, despite the detection of mucosal antibodies. Serum transfer experiments confirmed that circulating antibodies were able to migrate to the respiratory mucosa. Collectively, these results demonstrate that, especially upon repeated vaccination, the currently used COVID-19 mRNA vaccines can elicit mucosal neutralizing antibodies and that vaccination might also stimulate mucosal immunity induced by previous SARS-CoV-2 infection. Moreover, migration of circulating antibodies to the respiratory mucosa might be a main mechanism. These findings advance our understanding of mRNA vaccine-induced immunity and have implications for the design of vaccine strategies to combat respiratory infections.

Inter-laboratory harmonization of microsphere immunoassays for SARS-CoV-2 antibody detection in dried blood spots and oral fluids.

Dried blood spots (DBS) and oral fluids (OF) are easily attainable biospecimen types that have enabled population scale antibody monitoring for SARS-CoV-2 exposure and vaccination. However, the degree to which the two different biospecimen types can be used interchangeably remains unclear. To begin to address this question, we generated contrived DBS (cDBS) and OF (cOF) from serum panels from SARS-CoV-2 infected, vaccinated, and uninfected individuals. The contrived samples were evaluated using SARS-CoV-2 multiplexed microsphere immunoassays (MIAs) at two different institutions. Intra-laboratory tests revealed near perfect agreement between cDBS and cOF for N and S antigens, as evidenced by κ = 0.97-1 and 98%-100% agreement. Inter-laboratory comparisons were equally robust for both N (κ = 0.94-0.96; 97.5%-98 % agreement) and S (κ = 0.98 -1.0; 99.0%-100%). Furthermore, assays were transferred between labs, including methods and reagents, and a subset of cDBS and cOF samples (n = 52) were tested. Qualitative concordance remained high (κ = 0.94-1.0; 97.5%-100% agreement), confirming that integrity of the assays is retained upon transfer. In summary, our results provide evidence that DBS and OF can be used interchangeably across laboratories and institutions for the qualitative assessment of SARS-CoV-2 antibody determinations.

Detection of anti-leptospiral antibodies using recombinant ErpY-like lipoprotein based latex agglutination test for serodiagnosis of animal leptospirosis.

Precise and timely diagnosis is essential to prevent severe outcomes of leptospirosis in humans and animals. Existing diagnostic methods face challenges and limitations, underscoring the need for novel, field-applicable screening, and diagnostic tests/assays. This study evaluates the diagnostic utility of a recombinant ErpY-like lipoprotein (rErpY-LIC11966) in a latex agglutination test (LAT) for diagnosis of animal leptospirosis. The ErpY gene sequence from Leptospira interrogans serovar Pomona, excluding the signal peptide, was amplified, cloned into the pETite vector, and expressed in Escherichia coli. The expressed rErpY (∼16kDa) was characterized by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis and Western blot using Leptospira-specific standard sera. To assess the diagnostic potential of rErpY, Ni-NTA affinity-purified protein was used to sensitize latex-coated beads (0.8µm colour beads), which were then employed in the LAT for standardization and optimization with standard positive and negative sera. For evaluation, the rErpY-LAT was tested on serum samples from 177 suspected animal cases and compared to the microscopic agglutination test. It showed a relative diagnostic sensitivity of 90.6%, a specificity of 89.1%, and an overall accuracy of 90%. This study proposes rErpY-LAT as a field testing/screening diagnostic tool for preliminary serodiagnosis of leptospirosis, highlighting the potential of recombinant protein-based assays to address current diagnostic challenges.

Prevalence & clinical outcome of autoimmune encephalitis versus viral encephalitis in children with acute encephalitis syndrome: A prospective observational study.

Background & objectives Acute encephalitic syndrome (AES), encompasses a wide spectrum of potential causes, clinical presentations, and outcomes. While infectious encephalitis is generally considered more prevalent, autoimmune encephalitis is emerging as a significant aetiology. Neuronal autoantibodies have been identified independently or in association with acute viral encephalitis. The primary objective of this study was to ascertain the prevalence and clinical manifestation of autoimmune encephalitis as well as of coexisting viral markers in children with AES. Methods This study was a prospective observational investigation conducted in a hospital setting. It involved enrolling children with AES who were admitted to specific tertiary hospitals. Children were subjected to examinations to detect the presence of viral markers and neuronal autoantibodies in both their blood and cerebrospinal fluid (CSF). All the participants received treatment based on established guidelines and was followed for six months for outcome assessment. Results During the study period, 867 children with AES were examined. Among these cases, 37 children (4.2%) were diagnosed with autoimmune encephalitis, and all of them tested positive for anti-NMDAR (N-methyl-D-aspartate receptor) antibodies. Evidence of viral infection was seen in 409 (47.1%) of cases, out of which nearly 254 (29.2%) children had detectable HSV IgM antibodies. Among the 37 children with autoimmune encephalitis, 25 (67.5%) had evidence of a viral trigger, with eight of them tested positive for HSV IgM antibodies. The clinical presentation of autoimmune-associated AES was similar to those with viral aetiology. Interpretation & conclusions Autoimmune encephalitis triggered by neurotropic (HSV) viral infection was more prevalent in this study than in the earlier reports. Typically, these children show positive responses to immunosuppressive treatments if administered promptly. It is hence advisable to assess children who exhibit behavioural issues and movement disorders for possible autoimmune encephalitis.

Nationwide Seroprevalence of Dirofilaria immitis Antigen and Antibodies to Borrelia burgdorferi, Anaplasma spp., and Ehrlichia spp. in Shelter Cats in the United States, 2007–2011

Vector-borne infections persist as a significant issue in both human and animal health. Many of the most common vector-borne infections in the USA, especially tick-borne infections, are known to be zoonotic, including Lyme disease, anaplasmosis, and ehrlichiosis, and these infections may also negatively impact the health of infected animals. Convenient patient-side assays for the detection of antibodies to Borrelia burgdorferi, Anaplasma spp., and Ehrlichia spp., and antigen of Dirofilaria immitis have allowed for the generation of robust and large-scale prevalence data in dogs. Data of similar scale and distribution are not available in cats, and most feline prevalence studies have evaluated a small sample size with limited geographic distribution. The objective of this study was to evaluate the prevalence of antibodies to B. burgdorferi, Anaplasma spp., and Ehrlichia spp., and D. immitis antigen among shelter cats across the USA, a population that is presumably at high risk for ectoparasitism and, consequently, exposure to vector-borne infections. In total, 2232 whole blood samples were collected from shelter cats across four regions of the USA—South, Northeast, Midwest, and West—and were evaluated using the Idexx SNAP® 4Dx® Test. Ectoparasites were also opportunistically collected from cats during blood collection and morphologically identified. The prevalence of at least one vector-borne infection was 2.60%, and the nationwide prevalence was 1.88% for B. burgdorferi, 0.54% for Anaplasma spp., 0.09% for Ehrlichia spp., and 0.55% for D. immitis. A total of 1120 ectoparasites were collected from 423 cats, including 27 ticks and 1093 fleas. Although the overall prevalence of the pathogens in this survey is relatively low, we observe that there is an increased exposure risk regionally for some agents, with geographic distributions in this study mostly coinciding with established human and canine distributions. Understanding these findings in an assumed non-protected population of cats allows us to extrapolate the risk to pet cats if they are not provided routine veterinary care, including a broad-spectrum parasite prevention program.

Development and optimization of a double antigen sandwich ELISA detecting for Senecavirus A antibodies based on VP2 protein.

Senecavirus A (SVA) is an emerging viral pathogen that threatens the global swine industry significantly. The major clinical symptoms of SVA-infected animals are vesicular lesions, but various diseases can cause the same symptoms, which makes it difficult to distinguish SVA from other vesicular diseases clinically. The absence of an effective and safe vaccine necessitates the development of a simple, specific, and sensitive serological detection method for SVA antibodies. The VP2 protein of SVA, characterized by high immunogenicity and sequence conservatism, is an essential target for serological diagnosis. In this study, a double-antigen sandwich enzyme-linked immunosorbent assay [ELISA (DAgS-ELISA)] based on VP2 protein expressed by Escherichia coli was established for SVA antibody detection. With a cutoff value of 0.237, this assay demonstrated outstanding performance, showing high sensitivity and sharp specificity, which is manifested in the absence of cross-reaction with classical swine fever virus (CSFV), African swine fever virus (ASFV), pseudorabies virus (PRV), and porcine reproductive and respiratory syndrome virus (PRRSV), and foot-and-mouth disease virus (FMDV) serotype A and O. Additionally, the repeatability of the method is remarkable, as shown by the coefficients variation (CV) of both the intra- and inter-assay below 10%. By detecting 166 clinical sera, it was found that the kappa value of the DAgS-ELISA was 0.78 compared with that of the virus neutralization test (VNT), indicating a high level of consistency. In general, this method has high sensitivity, sharp specificity, remarkable repeatability, sound consistency, and low cost, making it a reliable and effective tool for detecting SVA antibodies.IMPORTANCESVA has rapidly become prevalent in many countries, and its outbreaks have threatened the global swine industry significantly. The major clinical symptoms of SVA-infected animals are vesicular lesions that are similar to other vesicular diseases, making it difficult to distinguish SVA. Currently, no commercial vaccines are available for SVA; therefore, effective diagnosis of SVA infection is vital for its prevention and control. In this study, VP2 protein of SVA was expressed by E. coli, and a double-antigen sandwich enzyme-linked immunosorbent assay [ELISA (DAgS-ELISA)] for SVA antibodies detection was successfully established based on the VP2 protein. The DAgS-ELISA has a high sensitivity, sharp specificity, remarkable repeatability, sound consistency, and low cost for detecting SVA antibodies. Therefore, the DAgS-ELISA established in this study may be a reliable and effective tool for detecting SVA antibodies and may be used to strengthen the monitoring and prevention of SVA epidemic in the long run.

Immuno- and expression analysis of Ehrlichia canis immunoreactive proteins.

Ehrlichia canis is the primary etiologic agent of canine monocytic ehrlichiosis, a serious and sometimes fatal hemorrhagic disease of dogs. Diagnosis of E. canis infection is often retrospectively confirmed by serologic detection of antibodies by immunofluorescent microscopy. Our laboratory previously identified numerous major immunoreactive proteins with species-specific linear antibody epitopes that are useful for immunodiagnosis of CME. More recently, we have defined the entire antibody-reactive immunome of E. canis, substantially increasing the number of major immunoreactive proteins known to exist. In this study, we analyzed and compared seven recently identified antibody reactive E. canis proteins with established diagnostic antigens including tandem repeat proteins TRP19, TRP36 and TRP140 and observed comparable immunoreactivity. Many of these proteins were conserved in different E. canis strains. Multiple linear antibody epitopes were mapped in a highly conserved TRP (Ecaj_0126), including within the tandem repeat domain. Temporal antibody responses were examined, and multiple proteins reacted with antibodies in sera as early as 21 days post experimental infection. Host-specific expression of the proteins was examined which revealed that some proteins exhibited higher expression in mammalian cells, while others in tick cells. This study has identified new immunodiagnostic candidates that exhibit different host expression patterns, information which may be useful for developing ultrasensitive immunodiagnostics and effective vaccines for CME.

Detection of Toxoplasma Gondii IgG Antibody in Patients Infected with Toxoplasma by ELISA

Aim the current study to diagnosis of the IgG antibody to Toxolasma gondii, serum samples isolated from patients humans and healthy reviewers to AL-Diwaniyah hospital and different laboratories. 90 samples were collected from human infected with Toxoplasmosis including 60 samples of patients and 30 samples of healthy (control). The results of this study showed that concentration of the IgG antibody in patients infected with T.gondii were 47.43 while the concentration in control were 8.14 and the percentage of infection in patients (24/60) (40%) and in control (4/30) (13.33%). The results showed that IgG antibody found in both patients and control but its concentration in patients was higher this indicates a high concentration of IgG antibody with toxoplasmosis.

The Role of Aspergillus Antibody Detection in Allergic Bronchopulmonary Aspergillosis

Allergic bronchopulmonary aspergillosis (ABPA) is an allergic lung disorder characterized by an exaggerated immune response to fungal allergens, particularly from Aspergillus fumigatus. ABPA leads to excessive mucus production and impaired mucociliary clearance. When A. fumigatus conidia are inhaled, they germinate, release exoproteases and other substances that further impede mucociliary clearance, and activate the immune response. The immune response is known by an exaggerated Th2 response to Aspergillus antigens, leading to production of IgE antibodies, eosinophilic infiltration of the lungs, and the formation of immune complexes. These immune complexes can deposit in the lung tissue, leading to airway inflammation, bronchiectasis, and fibrosis. This disease mainly occurs in individuals diagnosed with bronchial asthma and cystic fibrosis. Increased levels of A. fumigatus total IgE and specific IgE, IgG, and IgA antibodies, play a role in diagnosing patients with ABPA.

Acoustic pipette and biofunctional elastomeric microparticle system for rapid picomolar-level biomolecule detection in whole blood.

Most biosensing techniques require complex processing steps that generate prolonged workflows and introduce potential points of error. Here, we report an acoustic pipette to purify and label biomarkers in 70 minutes. A key aspect of this technology is the use of functional negative acoustic contrast particles (fNACPs), which display biorecognition motifs for the specific capture of biomarkers from whole blood. Because of their large size and compressibility, the fNACPs robustly trap along the pressure antinodes of a standing wave and separate from blood components in under 60 seconds with >99% efficiency. fNACPs are subsequently fluorescently labeled in the pipette and are analyzed by both a custom, portable fluorimeter and flow cytometer. We demonstrate the detection of anti-ovalbumin antibodies from blood at picomolar levels (35 to 60 pM) with integrated controls showing minimal nonspecific adsorption. Overall, this system offers a simple and versatile approach for the rapid, sensitive, and specific capture of biomolecules.

Evaluation of a multiplexed immunoassay for assessing long-term humoral immunity Orthopoxviruses

BackgroundThe 2022 Monkeypox virus (MPXV) global outbreak boosted development of multiple serological assays to aid understanding of Mpox immunology. ObjectivesThe study aimed to assess a multiplexed solid-phase electrochemiluminescence immunoassay (Meso Scale Discovery (MSD)) for simultaneous detection of antibodies against MPXV, including A35, E8 and M1 antigens, along with corresponding Vaccina Virus (VACV) homologues and demonstrate its accuracy in assessing antibody titres post-vaccination and infection. MethodsAssay performance was assessed for simultaneous detection of antibodies against MPXV and corresponding VACV antigens. Sensitivity and specificity were evaluated with paediatric negatives (n = 215), pre- and post-IMVANEX vaccinated (n = 80), and MPXV (Clade IIb, n = 39) infected serum samples. ResultsThe assay demonstrated high specificity (75.68 % (CI: 69.01–81.29) - 95.98 % (CI:92.54–97.87)) and sensitivity (62.11 % (CI:52.06–71.21) - 98.59 % (CI:92.44 %–99.93 %)) depending on the Orthopoxvirus antigen. Preferential binding was observed between MPXV-infected individuals and MPXV antigens, while vaccinated individuals exhibited increased binding to VACV antigens. These results highlight differential binding patterns between antigen homologues in related viruses. ConclusionOverall, this assay demonstrates high sensitivities in detecting antibodies for multiple relevant MPXV and VACV antigens post-infection and post-vaccination, indicating its utility in understanding immune responses to Orthopoxviruses in current and future outbreaks and evaluating the immunogenicity of new-generation Mpox-specific vaccinations.

A Multiplexed Digital Platform Enables Detection of Attomolar Protein Levels with Minimal Cross-Reactivity.

Protein-based biomarkers are essential for disease diagnostics, yet their low abundance in biofluids often presents significant detection challenges for traditional enzyme-linked immunosorbent assay (ELISA) techniques. While various ultrasensitive methods such as digital ELISA have improved sensitivity, multiplex assays still suffer from considerable cross-reactivities that can compromise result accuracies. To address this challenge, we have developed barcoded Molecular On-bead Signal Amplification for Individual Counting (barcoded MOSAIC), a multiplexed digital ELISA technology that markedly reduces cross-reactivity by pairing barcoded detection antibodies with specific bead types. This approach enables the simultaneous detection of eight analytes from less than 9 μL of blood, with sensitivities ranging from midpicomolar to low-attomolar levels and a collective dynamic range exceeding seven logs across multiple analytes within a single multiplex assay. Additionally, barcoded MOSAIC is compatible with standard immunoassay reagents and workflows, utilizing a rapid, automatable flow cytometric readout for quantification, which makes it a highly accessible benchtop platform that is readily adoptable by both research and clinical laboratories, setting the stage for future translation into point-of-care applications.