Specific Recognition of β-Galactofuranose-Containing Glycans of Synthetic Neoglycoproteins by Sera of Chronic Chagas Disease Patients

Chagas disease (CD) can be accurately diagnosed by detecting Trypanosoma cruzi in patients’ blood using polymerase chain reaction (PCR). However, parasite-derived biomarkers are of great interest for the serological diagnosis and early evaluation of chemotherapeutic efficacy when PCR may fail, owing to a blood parasite load below the method’s limit of detection. Previously, we focused on the detection of specific anti-α-galactopyranosyl (α-Gal) antibodies in chronic CD (CCD) patients elicited by α-Gal glycotopes copiously expressed on insect-derived and mammal-dwelling infective parasite stages. Nevertheless, these stages also abundantly express cell surface glycosylphosphatidylinositol (GPI)-anchored glycoproteins and glycoinositolphospholipids (GIPLs) bearing nonreducing terminal β-galactofuranosyl (β-Galf) residues, which are equally foreign to humans and, therefore, highly immunogenic. Here we report that CCD patients’ sera react specifically with synthetic β-Galf-containing glycans. We took a reversed immunoglycomics approach that entailed: (a) Synthesis of T. cruzi GIPL-derived Galfβ1,3Manpα-(CH2)3SH (glycan G29SH) and Galfβ1,3Manpα1,2-[Galfβ1,3]Manpα-(CH2)3SH (glycan G32SH); and (b) preparation of neoglycoproteins NGP29b and NGP32b, and their evaluation in a chemiluminescent immunoassay. Receiver-operating characteristic analysis revealed that NGP32b can distinguish CCD sera from sera of healthy individuals with 85.3% sensitivity and 100% specificity. This suggests that Galfβ1,3Manpα1,2-[Galfβ1,3]Manpα is an immunodominant glycotope and that NGP32b could potentially be used as a novel CCD biomarker.

Detection of Circulating VZV-Glycoprotein E-Specific Antibodies by Chemiluminescent Immunoassay (CLIA) for Varicella–Zoster Diagnosis

Varicella and herpes zoster are mild symptoms-associated diseases caused by varicella–zoster virus (VZV). They often cause severe complications (disseminated zoster), leading to death when diagnoses and treatment are delayed. However, most commercial VZV diagnostic tests have low sensitivity, and the most sensitive tests are unevenly available worldwide. Here, we developed and validated a highly sensitive VZV diagnostic kit based on the chemiluminescent immunoassay (CLIA) approach. VZV-glycoprotein E (gE) was used to develop a CLIA diagnostic approach for detecting VZV-specific IgA, IgG, and IgM. The kit was tested with 62 blood samples from 29 VZV-patients classified by standard ELISA into true-positive and equivocal groups and 453 blood samples from VZV-negative individuals. The diagnostic accuracy of the CLIA kit was evaluated by receiver-operating characteristic (ROC) analysis. The relationships of immunoglobulin-isotype levels between the two groups and with patient age ranges were analyzed. Overall, the developed CLIA-based diagnostic kit demonstrated the detection of VZV-specific immunoglobulin titers depending on sample dilution. From the ELISA-based true-positive patient samples, the diagnostic approach showed sensitivities of 95.2%, 95.2%, and 97.6% and specificities of 98.0%, 100%, and 98.9% for the detection of VZV-gE-specific IgA, IgG, and IgM, respectively. Combining IgM to IgG and IgA detection improved diagnostic accuracy. Comparative analyses on diagnosing patients with equivocal results displaying very low immunoglobulin titers revealed that the CLIA-based diagnostic approach is overall more sensitive than ELISA. In the presence of typical VZV symptoms, CLIA-based detection of high titer of IgM and low titer of IgA/IgG suggested the equivocal patients experienced primary VZV infection. Furthermore, while no difference in IgA/IgG level was found regarding patient age, IgM level was significantly higher in young adults. The CLIA approach-based detection kit for diagnosing VZV-gE-specific IgA, IgG, and IgM is simple, suitable for high-throughput routine analysis situations, and provides enhanced specificity compared to ELISA.

Systemic COVID-19 vaccination also enhances the humoral immune response after SARS CoV-2 infection in the population of an oncology hospital in Poland. Criteria for COVID-19 re-immunization are needed.

Systemic vaccination of the BNT162b2 mRNA stimulates humoral response. Our study aimed to compare the intensity of humoral immune response, measured by SARS CoV-2 IgG, SARS CoV-2 IgM, and neutralization S-RBD IgG antibodies level, post COVID-19 vaccination versus post-SARS COV-2 infection. We analysed 1060 people in the following groups: convalescents, healthy vaccinated, vaccinated with COMIRNATY, AstraZeneca, Moderna, Johnson & Johnson, and vaccinated SARS CoV-2 convalescents. A concentration of SARS CoV-2 IgG, SARS CoV-2 IgM, and neutralizing S-RBD IgG was estimated in hospital laboratory by chemiluminescent immunoassay - CLIA, MAGLUMI. Results: 1. We observed a rise of antibodies response in both convalescent SARS CoV-2 and COVID-19 vaccinated groups 2. The level of all antibodies’ concentrations in vaccinated COVID-19 convalescents was significantly higher. 3. We differentiated asymptomatic SARS CoV-2 convalescents from the control group. Based on our analysis, we suggest that it is essential to monitor SARS CoV-2 antibodies concentrations as an indicator of asymptomatic COVID-19 infection and equivalent to the effectiveness of humoral response in convalescents and vaccinated people. Considering the time-limited nature of the effects of post-infection SARS CoV-2 recovery or vaccination, among others physiological half-life, we suggested monitoring IgG antibodies level as a criterium for the next vaccination.

Validity of Chemiluminescent Immunoassay Serology Test for Anti-Sars Cov-2 Antibodies IgM and IgG 1

In December 2019, an outbreak of acute pneumonia occurred in Wuhan, China. The disease was transmitted betweenhumans through droplets (coughing or sneezing) of infected patients, causing this outbreak to spread rapidly in variouscountries in the world, including Indonesia. On February 11, 2020, WHO announced the pneumonia was caused byCoronavirus Disease 2019 (COVID-19), which was caused by a new type of Coronavirus, the SARS-CoV-2. A rapid andaccurate diagnosis is critical for the control of the COVID-19 outbreak. The widely used test is a serology-based test thatdetects the presence of SARS-CoV-2 IgM/IgG antibodies in the patient's body. One of the methods used for this test isChemiluminescent Immunoassay (CLIA). This study aimed to determine the reliability of CLIA. The study was conductedfrom August to September 2020. The number of samples was 63 patients' serum. Polymerase chain reaction examination atHusada Utama Hospital, Surabaya, revealed that 21 patients were confirmed positive for COVID-19 with positive PCRresults, and 42 patients were healthy with negative COVID-19 results. The results showed that IgM had a diagnosticsensitivity of 85.7%, diagnostic specificity of 92.8%, a positive predictive value of 85.7%, a negative predictive value of 92.8%,and accuracy of 90.4%. In comparison, IgG had a diagnostic sensitivity of 90.4%, diagnostic specificity of 90.4%, a positivepredictive value of 82.6%, a negative predictive value of 90.5%, and accuracy of 90.4%. In conclusion, IgG has a highersensitivity than IgM, while IgM had higher specificity, positive predictive value, and negative predictive value than IgG.However, the positive, negative predictive value and efficiency values were the same for IgM and IgG.

Neuron-specific enolase level is a useful biomarker for distinguishing amyotrophic lateral sclerosis from cervical spondylotic myelopathy

The current study aimed to evaluate whether cerebrospinal fluid (CSF) neuron-specific enolase (NSE) levels are elevated in amyotrophic lateral sclerosis (ALS) and are effective in distinguishing ALS from cervical spondylotic myelopathy (CSM). We retrospectively evaluated 45 patients with ALS, 23 with CSM, 28 controls, and 10 with Parkinson’s disease (PD) who underwent analysis of CSF NSE levels. The control group comprised patients aged above 45 years who underwent lumbar puncture because of suspected neurological disorders that were ruled out after extensive investigations. CSF NSE levels were evaluated using the electro-chemiluminescent immunoassay. The ALS group had significantly higher CSF NSE levels than the CSM and control groups (P < 0.001 for both comparisons). The CSM, control, and PD groups did not significantly differ in terms of CSF NSE levels. A receiver-operating characteristic curve analysis was performed to assess the diagnostic value of CSF NSE levels in distinguishing ALS from CSM. The area under the curve for CSF NSE levels was 0.86. The optimal cutoff value was 17.7 ng/mL, with a specificity of 87% and a sensitivity of 80%. Hence, CSF NSE levels are elevated in ALS and are effective in distinguishing ALS from CSM.

Preparation of Anti-Aristolochic Acid I Monoclonal Antibody and Development of Chemiluminescent Immunoassay and Carbon Dot-Based Fluoroimmunoassay for Sensitive Detection of Aristolochic Acid I

Aristolochic acid (AA) toxicity has been shown in humans regarding carcinogenesis, nephrotoxicity, and mutagenicity. Monitoring the AA content in drug homologous and healthy foods is necessary for the health of humans. In this study, a monoclonal antibody (mAb) with high sensitivity for aristolochic acid I (AA-I) was prepared. Based on the obtained mAb, a chemiluminescent immunoassay (CLEIA) against AA-I was developed, which showed the 50% decrease in the RLUmax (IC50) value of 1.8 ng/mL and the limit of detection (LOD) of 0.4 ng/mL. Carbon dots with red emission at 620 nm, namely rCDs, were synthesized and employed in conventional indirect competitive enzyme-linked immunosorbent assay (icELISA) to improve the assay sensitivity of a fluoroimmunoassay (FIA). Oxidized 3,3′′,5,5′′-tetramethylbenzidine dihydrochloride (oxTMB) can quench the emission of the rCDs through the inner-filter effect; therefore, the fluorescence intensity of rCDs can be regulated by the concentration of mAb. As a result, the assay sensitivity of FIA was improved by five-fold compared to CLEIA. A good relationship between the results of the proposed assays and the standard ultra-high performance liquid chromatography-triple quadrupole mass spectrometer (UPLC-QQQ-MS/MS) of real samples indicated good accuracy and practicability of CLEIA and FIA.

Systemic COVID-19 vaccination also enhances the humoral immune response after SARS CoV-2 infection in the population of an oncology hospital in Poland. Criteria for COVID-19 re-immunization are needed.

Systemic vaccination of the BNT162b2 mRNA stimulates humoral response. The aim of our study was to compare the intensity of humoral immune response, measured by SARS CoV-2 IgG, SARS CoV-2 IgM, and neutralization S-RBD IgG antibodies level, post COVID-19 vaccination versus post SARS COV-2 infection. We analysed 1060 people in the following groups: convalescents, healthy vaccinated, vaccinated with COMIRNATY, AstraZeneca, Moderna, Johnson&Johnson and vaccinated SARS CoV-2 convalescents. A concentration of SARS CoV-2 IgG, SARS CoV-2 IgM, and neutralizing S-RBD IgG was estimated in Bialystok Oncology Center laboratory by chemiluminescent immunoassay- CLIA, MAGLUMI. Results: 1. We observed a raise of antibodies response in both, convalescent SARS CoV-2 and COVID-19 vaccinated groups 2. The level of all antibodies’ concentrations in vaccinated COVID-19 convalescents was significantly higher. 3. We differentiated an asymptomatic SARS CoV-2 convalescents from control group. Based on our analysis we suggest that it is important to monitor SARS CoV-2 antibodies concentrations as an indicator of asymptomatic COVID-19 infection, and as an equivalent of effectiveness of humoral response in convalescents and vaccinated people. Taking into consideration the time-limited nature of the effects of post infection SARS CoV-2 recovery or vaccination, among others physiological half-life, we suggested monitoring IgG antibodies level as a criterium for next vaccination.

Comparisons of Anti-dsDNA Antibody Detection Methods by Chemiluminescent Immunoassay and Enzyme-Linked Immunosorbent Assay in Systemic Lupus Erythematosus

This study aimed to compare the test results of anti-double-stranded DNA (anti-dsDNA) antibodies obtained using chemiluminescent immunoassay (CIA) and enzyme-linked immunosorbent assay (ELISA), and investigate predictors of inconsistent results. This retrospective study included 502 patients who underwent CIA and ELISA to determine their anti-dsDNA antibody values within a year. We compared the diagnostic power for SLE, disease activity, and predictive power for lupus nephritis (LN). A multivariate analysis was performed to determine the predictors of inconsistencies. CIA and ELISA were moderately correlated in terms of their consistency (Cronbach’s α = 0.571), and yielded comparably favorable results in terms of SLE diagnostic power and SLE disease activity. However, if the patient had LN, CIA displayed higher predictive power than ELISA (0.620 vs. 0.555, p = 0.026). Compared with the CIA/ELISA double-positive group, the inconsistent group had lower anti-C1q circulating immune complexes (CIC) antibody values (OR: 0.42, 95% CI: 0.18–0.94, p = 0.036), and lower SLEDAI scores (≥4) (OR: 0.33, 95% CI: 0.14–0.79, p = 0.013). Anti-dsDNA antibody detection with CIA exhibited higher predictability for diagnosing LN than did ELISA. In the event of inconsistencies between anti-dsDNA methods, SLE disease activity and CIC test values should be considered simultaneously.