Automated Chemiluminescence Immunoassay Research Articles

Specific reference interval for high-sensitivity cardiac troponin I among healthy children in Wuhan, China.

Troponin (Tn) is of an important biomarker for the diagnosis and prognosis of myocardial injury and for evaluating the severity of cardiac involvement due to other systemic diseases in children. Unfortunately, high-sensitivity cardiac troponin I (hs-cTnI) specific reference intervals (RIs) are extremely limited. This study aimed to establish a preliminary pediatric hs-cTnI RI for newborns, children, and adolescents in Wuhan, China. A total of 1,355 healthy participants (1 day to 19 years) were recruited from a cross-sectional study implemented in Wuhan Children's Hospital from September 2022 to August 2023. Serum hs-cTnI levels were detected via the Mindray automated chemiluminescence immunoassay analyzer (CL-6000i). Specific serum hs-cTnI RIs were established according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. The RIs were defined by the nonparametric median (P50), and 2.5th, 97.5th [P50 (P2.5-P97.5)] intervals. Of the 1,355 pediatric participants, serum hs-cTnI concentrations of 1,332 children were measured. The serum overall P50 and 95% interval range (P2.5-P97.5) of serum hs-cTnI was 0.41 (0.00, 44.31) ng/L. This was higher in males of 0.47 (0.00, 44.90) ng/L than in females of 0.36 (0.00, 44.17) ng/L (P<0.01). Age- and sex-specific differences in hs-cTnI levels were observed. The levels were highly variable in children under 1 year of age (especially in newborns), deriving a P50 (P2.5-P97.5) of 22.06 (1.04, 154.22) ng/L, and gradually narrowed and decreased with increasing age, with a markedly lower established P50 (P2.5-P97.5) of 0.36 (0.00, 2.16) ng/L. However, the levels began to increase slightly at the age of 9-12 years and reached a small peak at the age range of 15 to 18 years in males with 0.71 (0.03, 3.29) ng/L, while females were less affected by puberty. Sex- and age-specific RIs for hs-cTnI were established: 5 age-specific RIs in males, 1 day-1 month: 30.16 (8.67, 171.81) ng/L; >1-12 months: 13.20 (0.63, 61.91) ng/L; >1-15 years: 0.36 (0.00, 1.86) ng/L; >15-18 years: 0.71 (0.03, 3.29) ng/L; >18-19 years: 0.52 (0.00, 1.92) ng/L; and 4 age-specific RIs in females, 1 day-1 month: 43.93 (18.82, 146.38) ng/L; >1-12 months: 5.22 (0.92, 42.54) ng/L; >1-6 years: 0.54 (0.00, 2.74) ng/L; >6-19 years: 0.23 (0.00, 1.56) ng/L. This study preliminarily established age- and sex-specific serum hs-cTnI RIs using the Mindray CL-6000i system in healthy children in Wuhan, China.

Assessment of the Diagnostic Performance of Fully Automated Hepatitis E Virus (HEV) Antibody Tests.

The detection of anti-hepatitis E virus (HEV) antibodies contributes to the diagnosis of hepatitis E. The diagnostic suitability of two automated chemiluminescence immunoassays (CLIAs, LIAISON® MUREX Anti-HEV IgG/Anti-HEV IgM test, DiaSorin) was assessed by comparison with the results of a combination of enzyme immunoassays and immunoblots (recomWell HEV IgG/IgM ELISA, recomLine HEV IgG/IgM, MIKROGEN). Samples with a deviating result were analyzed with the WANTAI ELISAs. Compared to the recomWell ELISAs, the Anti-HEV IgG CLIA had a percentage overall agreement (POA) of 100% (149/149; 95% CI: 97.5-100%) and the Anti-HEV IgM CLIA had a POA of 83.3% (85/102; 95% CI: 74.9-89.3%); considering the recomLine HEV IgM results, the POA was 71.7% (38/53; 95% CI: 58.4-82%). The WANTAI test confirmed 52.9% (9/17) of negative CLIA IgMs; HEV RNA was not detectable. Since acute infection with the Epstein-Barr virus (EBV) or human cytomegalovirus (CMV) may influence the results of other serological assays, HEV antibodies were examined in 17 EBV and 2 CMV patients: One had an isolated and probably unspecific HEV IgM in the CLIA, as HEV RNA was not detectable. Both CLIAs are well suited for HEV diagnostics, but isolated IgM should be confirmed. An acute EBV/CMV infection can influence HEV serodiagnostics.

Serum apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC) is a novel stroke biomarker

BackgroundApoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC) is a promising stroke biomarker. However, a large study of human serum ASC has not yet to be reported; additionally, the diagnostic value of prehospital concentration and practicality of ASC remains unknown. MethodsWe recruited 774 Chinese stroke patients, including 523 with ischemic stroke (IS) and 251 with hemorrhagic stroke (HS) within 14 days following symptom onset in the emergency department, alongside 481 healthy individuals and 64 cognitive impairment patients as controls. Serum ASC concentrations were determined using automated chemiluminescence immunoassay, exploring the relationship between serum ASC concentration and subtypes, severity, and sampling timepoints of stroke. ResultsASC concentrations were significantly higher in stroke patients compared with all controls (P < 0.001). HS patients had greater ASC concentrations than IS patients (P < 0.05). With increasing ASC concentration, the proportion of severe cases increased. The area under the receiver operating characteristic curve (AUC) for differentiating between healthy individuals and stroke patients in the hyperacute phase was 0.78; this markedly improved (0.90) when considering samples from healthy individuals and patients with subarachnoid hemorrhage (SAH) ≤ 3 h from last-known-well (LKW). ConclusionsSerum ASC is a valuable biomarker for stroke differentiation and aids in the clinical diagnosis of stroke severity and subtypes.

Serum KL-6 as a Candidate Predictor of Outcome in Patients with SARS-CoV-2 Pneumonia.

Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2)-infection is associated with an extremely variable disease course. When interstitial pneumonia (IP) occurs, it can lead to acute respiratory distress syndrome and death. Serum Krebs von den Lungen-6 (KL-6) is an established marker of IP, but its role as a marker of SARS-CoV-2 pneumonia is debated. This bicentric study included 157 patients with SARS-CoV-2 pneumonia. The WHO Ordinal Scale for Clinical Improvement (0-10 points) was used to classify the clinical course. Serum samples were collected at admission, and on days 3 and 7 of hospitalization. KL-6 was measured by using automated chemiluminescence immunoassay. A total of 68 patients developed a severe SARS-CoV-2 pneumonia, 135 of them required oxygen, and 15 died during hospitalization. The patients requiring non-invasive ventilation, invasive ventilation, or extracorporeal membrane oxygenation had significantly higher serum KL-6 levels at admission. The serum KL-6 levels were tendentially higher in patients who died than in those who survived. Logistic regression identified serum KL-6 at a cut-off of 335 U/mL at admission as a significant predictor of severe SARS-CoV-2 pneumonia outcome. Serum KL-6 seems to be a candidate biomarker for the clinical routine to stratify patients with SARS-CoV-2 pneumonia for the risk of a severe disease outcome or death.

Hepatitis E Prevalence in Vulnerable Populations in Goiânia, Central Brazil.

A transversal study was conducted among 472 vulnerable individuals (recyclable waste pickers, immigrants and refugees, homeless individuals, as well as lesbian, gay, bisexual, and transexual individuals) in Goiânia City, the capital of the State of Goiás, Brazil, to investigate the prevalence of hepatitis E virus (HEV) infection. A total of 459 (97.2%) serum samples were tested for anti-HEV IgG and IgM antibodies using fully automated chemiluminescence immunoassays (Liaison® Murex Anti-HEV IgG and IgM assays, DiaSorin, Saluggia, Italy). Positive samples were tested for the presence of HEV RNA by a real-time polymerase chain reaction. A seroprevalence of 0.87% (95% confidence interval [CI]: 0.34-2.22) was found for anti-HEV IgG. Furthermore, anti-HEV IgM was detected in only one individual (0.22%; 95% CI: 0.04-1.22), who was also negative for HEV RNA. These findings revealed that HEV infection is infrequent in vulnerable individuals in Central Brazil, with low seroprevalence of past and recent HEV infections.

B-058 Development and Evaluation of the Assay for Detection of Human Specific IgE Against Animal Dander Allergens on a Fully Automated Chemiluminescence Immunoassay Platform—Autolumo® A2000 Plus

Abstract Background The detection of specific IgE is a key method for diagnosing allergic diseases. An accurate, user friendly and cost-effective way to clinically diagnosing allergies is beneficial for the patients. Our detection kit together with the Autolumo A2000 Plus chemiluminescence fully automated platform provide a new and more effective (40 minutes) way for detection of specific IgE with simple operation, high sensitivity and specificity. Methods The Animal Dander Allergen Mix Assay kit detects specific IgE against cat dander (E1), horse dander (E3), cow dander (E4) and dog dander (E5). Performance evaluation of the animal dander mix, E1, E3, E4 and E5 assay kits for the detection of specific IgE in serum was performed on our automated chemiluminescence platform (Autolumo A2000 Plus) and compared to a well-characterized platform (ImmunoCAP, Phadia). During detection, magnetic particle suspension, antigen solution, patient sample, and enzyme conjugate were added sequentially. A streptavidin-biotinylated antigen-sIgE antibody-anti-IgE-HRP conjugate complex is formed, wherein the amount of the anti-IgE-HRP conjugate is positively correlated to the amount of sIgE antibodies in the patient sample. After the substrate is added, the HRP catalyze the chemical reaction to generate a recordable signal in the form of light. The intensities of the light were obtained by using a chemiluminescence quantitative analyzer, and the standard curve was generated according to the 7-point calibration, and then the sIgE antibody concentrations in the serum sample were analyzed. The overall agreement, method comparison, precision, linearity, and stability of our mixed and single assay kits were evaluated using serum samples (n &amp;gt; 140) that had been characterized with Phadia. Results The overall agreement for the animal dander mix assay is 91.84%, with the average intra-run and inter-run precision to be 1.97% and 4.21% respectively. The limit of blank (LoB) and limit of detection (LoD) for the animal dander mix assay is 0.019 kU/L and 0.045 kU/L respectively. The sensitivity for our single E1, E3, E4, and E5 assays are 97%, 85.6%, 70.8%, and 80.2%, respectively. All single assays have a specificity &amp;gt;90%, total precision &amp;lt;5% CV, and a linearity with R2 &amp;gt; 0.999. The LoB, LoD and recommended LoQ for all single assays are 0.019 kU/L, 0.037 kU/L, and 0.1 kU/L respectively. The average signal drifts after 10 days incubation in 37C is less than 10% indicating the assays are stable. Conclusion Autolumo A2000 Plus is a fully automated chemiluminescence Immunoassay platform that provides accurate and cost-effective screening for allergy diagnostics with excellent overall agreement, precision, and stability.

Distribution characteristics of plasma renin concentration in patients with aldosterone-producing adenoma

Objective: To analyze the distribution characteristics of plasma renin concentration (PRC) in patients with aldosterone-producing adenoma (APA) and its impact on diagnosis. Methods: In this retrospective case series, clinical data from 200 patients with APA (80 men and 120 women; mean age 45.6 years) in the First Affiliated Hospital of Chongqing Medical University from November 2013 to January 2022 were evaluated. PRC was determined by automated chemiluminescence immunoassay. The distribution characteristics of PRC were analyzed, and 8.2 mU/L was used as the low renin cutoff to evaluate whether renin was suppressed. Results: The median PRC was 1.6 mU/L (range, 0.4-41.5 mU/L). There were 116 patients with APA with PRC of ≤2 mU/L, 41 patients with 2<PRC≤4 mU/L. PRC was not suppressed (PRC>8.2 mU/L) in 8.0% (16/200) of the patients with APA. And PRC was not suppressed in 2.5% (5/200) of the patients with APA, resulting in a primary aldosteronism negative screening outcome. Conclusions: Although most patients with APA have low PRC, there are a small number (8%) of patients whose PRC has not been fully suppressed, which can lead to missed diagnoses during primary aldosteronism screening. While primary aldosteronism is highly suspected, further investigations are required to determine the diagnosis, even if PRC is not fully suppressed at screening.

Retrospective Multicenter Evaluation of the VirClia Galactomannan Antigen Assay for the Diagnosis of Pulmonary Aspergillosis with Bronchoalveolar Lavage Fluid Samples from Patients with Hematological Disease.

Galactomannan (GM) testing of bronchoalveolar lavage (BAL) fluid samples has become an essential tool to diagnose invasive pulmonary aspergillosis (IPA) and is part of diagnostic guidelines. Enzyme-linked immunosorbent assays (ELISAs) (enzyme immunoassays [EIAs]) are commonly used, but they have a long turnaround time. In this study, we evaluated the performance of an automated chemiluminescence immunoassay (CLIA) with BAL fluid samples. This was a multicenter retrospective study in the Netherlands and Belgium. BAL fluid samples were collected from patients with underlying hematological diseases with a suspected invasive fungal infection. Diagnosis of IPA was based on the 2020 European Organisation for Research and Treatment of Cancer (EORTC)/Mycoses Study Group Education and Research Consortium (MSGERC) consensus definitions. GM results were reported as optical density index (ODI) values. ODI cutoff values for positive results that were evaluated were 0.5, 0.8, and 1.0 for the EIA and 0.16, 0.18, and 0.20 for the CLIA. Probable IPA cases were compared with two control groups, one with no evidence of IPA and another with no IPA or possible IPA. Qualitative agreement was analyzed using Cohen's κ, and quantitative agreement was analyzed by Spearman's correlation. We analyzed 141 BAL fluid samples from 141 patients; 66 patients (47%) had probable IPA, and 56 cases remained probable IPA when the EIA GM result was excluded as a criterion, because they also had positive culture and/or duplicate positive PCR results. Sixty-three patients (45%) had possible IPA and 12 (8%) had no IPA. The sensitivity and specificity of the two tests were quite comparable, and the overall qualitative agreement between EIA and CLIA results was 81 to 89%. The correlation of the actual CLIA and EIA values was strong at 0.72 (95% confidence interval, 0.63 to 0.80). CLIA has similar performance, compared to the gold-standard EIA, with the benefits of faster turnaround because batching is not required. Therefore, CLIA can be used as an alternative GM assay for BAL fluid samples.

Evaluation of the Chagas VirClia® and Chagas TESA VirClia® for the Diagnosis of Trypanosoma cruzi Infection.

Chagas disease (CD), caused by the protozoan Trypanosoma cruzi, is an important problem of public health even in regions where it is not endemic. Spain ranks second worldwide in terms of imported cases of T. cruzi infection in the chronic phase. The diagnosis in this stage is made via the detection of antibodies against T. cruzi. Therefore, we aimed to evaluate the sensitivity and specificity of two fully automated chemiluminescence immunoassays, Chagas VirClia® (CHR), which uses a mixture of recombinant antigens, and Chagas TESA VirClia® (TESA), the first chemiluminescence assay based on excretion-secretion antigens of trypomastigotes, both designed in monotest format. A retrospective case-control study was performed using 105 well-characterized samples: 49 from patients with CD, 22 from uninfected individuals, and 32 from patients with other pathologies. Sensitivity was 98% for CHR and 92% for TESA. In contrast, the specificity in both was 100%. Cross-reactivity was observed in leishmaniasis (2/10). CHR meets the criteria to become a tool for serological screening, while TESA has the potential for confirmation and cross-reaction discrimination. The monotest format allows its application in laboratories with a small number of samples. The high specificity of both assays is useful in areas where leishmaniasis is endemic.

Screening for maternal and congenital syphilis with a chemiluminescence immunoassay in a South African private specialist healthcare sector setting

&#x0D; &#x0D; &#x0D; &#x0D; Background. Syphilis is a sexually transmitted infection that is most frequently found in lower socio-economic groups globally and is associated with significant maternal and fetal complications. In South Africa (SA), the last two to three decades have seen a rise in the number of people in the low and middle economic social groups seeking private specialist healthcare services.Objective. To evaluate the prevalence rates of maternal and congenital syphilis in a private specialist healthcare setting.&#x0D; Methods. The laboratory case records of women who had antenatal maternal syphilis (MS) screening using the automated chemiluminescence immunoassay (Architect Syphilis TP) in a private laboratory facility in Durban were reviewed.Results. A total of 9 740 individual maternal serum samples were analysed and 256 were Architect Syphilis TP positive, resulting in a MS prevalence rate of 2.7%. Of the less than three-quarters of exposed neonates tested (71.1%; n=182/256), 38.5% (n=70/182) were Architect syphilis TP positive. Less than a tenth of exposed neonates (2.43%; n=6) had only rapid plasma reagin (RPR) titers test whereas 26.6% (n=68/256) did not have a syphilis screen test. Based on the 182 exposed neonates tested, the congenital syphilis (CS) prevalence from the laboratory records was 7.7%. The highest rate of MS was in the ≥35 years age group.&#x0D; Conclusion. The prevalence of MS in the private specialist healthcare sector in SA is relatively high and warrants continued maternal antenatal screening during early pregnancy across all socio-economic groups. The high rate of MS in the age group over 35 years warrants further investigations and explanation.&#x0D; &#x0D; &#x0D; &#x0D;

Relationship between inflammatory biomarkers, vitamin D levels, and depressive symptoms in late pregnancy and during the postpartum period: a prospective, observational study

BackgroundPerinatal depression impacts maternal and fetal health, and exhibits a high rate of continuity postpartum. Not only does it impair the maternal quality of life, it also increases the risk of adverse birth and developmental problems in offspring. Vitamin D deficiency and excessive inflammation have been associated with perinatal depression. There is a scarcity of evidence regarding the biological causes of maternal depression in Iraq, therefore, the present study aims to assess perinatal depressive symptoms associations with inflammatory markers and vitamin D levels, and to investigate the interaction between vitamin D and the inflammatory markers. A prospective, observational study design was utilized to recruit healthy pregnant women from private obstetrics clinic in Baghdad, Iraq, from April to September 2021. The Edinburgh Postnatal Depression Scale (EPDS) was used to measure depressive symptoms during the third trimester and at 6 months postpartum. Serum levels of interleukin (IL)-6, C-reactive protein (CRP), and 25-hydroxy vitamin D (25-OH-D) were quantified, using a fully automated chemiluminescence immunoassay analyzer.ResultsEighty patients were eligible for inclusion. The antenatal EPDS scores demonstrated a significant association with square root IL-6 (B = – 0.025, p = 0.040) and no association with CRP or vitamin D levels. The severity of postpartum depressive symptoms tended towards a positive association, with larger increases of CRP concentration (p = 0.065). In contrast, the association between marital relationship quality and CRP was statistically significant (p = 0.001). There was a statistically significant association between CRP and vitamin D concentration (p = 0.041). Antepartum EPDS significantly predicted the postpartum EPDS score (p = 0.000, B = 0.180, R2 for the model = 0.976, CI (0.17–0.19)).ConclusionsThe study findings show a significant association between third trimester depressive symptoms and IL-6 concentration. CRP and vitamin D levels do not correlate with perinatal depressive symptoms and a poor marital relationship significantly elevates the CRP level. In addition, vitamin D level was associated with CRP level and antepartum depressive symptoms predict postpartum EPDS score. Future studies involving a larger population and including women with pregnancy complications would provide a further insight into the role of inflammation and vitamin D deficiency in the etiology of perinatal depression.

The effects of combined amplitude and high-frequency vibration on physically inactive osteopenic postmenopausal women.

Purpose: To evaluate whole-body vibration (WBV) osteogenic potential in physically inactive postmenopausal women using high-frequency and combined amplitude stimuli. Methods: Two-hundred fifty-five physically inactive postmenopausal women (55–75 years) with 10-year major osteoporotic fracture risk (3%–35%) participated in this 18-month study. For the first 12 months, the vibration group experienced progressive 20-min WBV sessions (up to 3 sessions/week) with rest periods (30–60 s) between exercises. Frequencies (30–50 Hz), with low (0.2–0.4 mm) and high (0.6–0.8 mm) amplitude stimuli were delivered via PowerPlate Pro5 platforms producing accelerations of (0.75–7.04 g). The last 6 months for the treatment group were a follow-up period similar to control. Serum bone remodelling markers [C-terminal crosslinked telopeptide of type-1 collagen (CTX), procollagen type-1 N-terminal propeptide (P1NP), bone alkaline phosphatase (BAP) and sclerostin] were measured at fasting. CTX and P1NP were determined by automated chemiluminescence immunoassay, bone alkaline phosphatase (BAP) by automated spectrophotometric immunoassay, and sclerostin by an enzyme-immunoassay. Bone mineral density (BMD) of the whole-body, proximal femur and lumbar vertebrae was measured by dual-energy X-ray absorptiometry (DXA). Bone microarchitecture of the distal non-dominant radius and tibia was measured by high-resolution peripheral quantitative computed tomography (HR-pQCT). Results: Femoral neck (p = 0.520) and spine BMD (p = 0.444) failed to improve after 12 months of WBV. Bone macro and microstructural parameters were not impacted by WBV, as well as estimated failure load at the distal radius (p = 0.354) and tibia (p = 0.813). As expected, most DXA and HR-pQCT parameters displayed age-related degradation in this postmenopausal population. BAP and CTX increased over time in both groups, with CTX more marginally elevated in the vibration group when comparing baseline changes to month-12 (480.80 pmol/L; p = 0.039) and month-18 (492.78 pmol/L; p = 0.075). However, no differences were found when comparing group concentrations only at month-12 (506.35 pmol/L; p = 0.415) and month-18 (518.33 pmol/L; p = 0.480), indicating differences below the threshold of clinical significance. Overall, HR-pQCT, DXA bone parameters and bone turnover markers remained unaffected. Conclusion: Combined amplitude and high-frequency training for one year had no ameliorating effect on DXA and HR-pQCT bone parameters in physically inactive postmenopausal women. Serum analysis did not display any significant improvement in formation and resorption markers and also failed to alter sclerostin concentrations between groups.

Performance evaluation of eight treponemal antibody tests in China

In this study, the performance of 8 commercially available treponemal antibody tests was evaluated. Higher specificity was achieved in the automated chemiluminescence immunoassays(CIA assays) than enzyme-linked immunosorbent (ELISA) assays, but no significant differences were found in terms of the sensitivity levels. The levels of test sensitivity did not differ by HIV status and TRUST titer but did differ by clinical stage. The results of WB-IgG revealed that the level of antibodies against the 4 antigens showed an upward trend with the development of the clinical stages of syphilis, reaching a peak in tertiary syphilis and then decreasing. The present results supported the good performance of CIA tests used in the blood source screening of syphilis, which reduced the waste of plasma. The reaction intensities of anti-TP17 and TP47 antibodies in past treated syphilis samples were considerably weak and were expected to be used for monitoring the therapeutic effect of syphilis.

Vitamin D Deficiency and Risk Factors Related to Acute Psychiatric Relapses in Patients with Severe Mental Disorders: A Preliminary Study.

Previous studies have indicated that vitamin (Vit) D deficiency is frequent in psychiatric patients, regardless of diagnostic category. We aimed to assess whether acute psychiatric relapses in inpatients was associated with Vit D deficiency compared to stabilized outpatients. The cohort (152 total patients, 75 males and 77 females) had a mean age of 47.3 ± 14.4 years at admission and was grouped according to psychiatric diagnosis. Psychopathological symptom severity was assessed by the Brief Psychiatric Rating Scale (BPRS), a multidimensional symptom inventory. Total calcium serum levels were measured using standard laboratory methods, while plasma levels of 25-OH-Vit D and parathyroid hormone (PTH) were measured by automated chemiluminescence immunoassays. The psychiatric inpatient subgroup showed a significant difference in serum levels of 25-OH-Vit D and PTH (p < 0.001). Correlation analysis between serum levels of 25-OH-Vit D and BPRS total and subitem scores indicated a significantly negative relationship. In addition, linear regression analysis evidenced that the inpatient condition might predict low PTH and 25-OH-Vit D serum levels. Hospitalized psychiatric patients are at increased risk for Vit D deficiency regardless of their diagnostic categories. The mechanism underlying the association between acute psychiatric relapses and Vit D deficiency remains unclear. Therefore, screening for Vit D deficiency should pertain to the health assessment of patients with major psychiatric disorders.

Development of a self-assembled heptameric nanobody/streptavidin-binding peptide fusion for ultrasensitive detection of serum biomarkers

Since serum biomarkers are commonly used as the indicators of human health, sensitive detection methods for biomarkers can facilitate the diagnosis of related diseases. Herein, to improve sensitivity of the traditional enzyme immunoassay for the model macromolecule alpha fetoprotein (AFP), a novel signal amplification strategy using nanobody/streptavidin-binding peptide (Nb/SBP) heptamer and streptavidin poly-horseradish peroxide conjugate (SA-polyHRP) was proposed to develop an Nb/SBP heptamer-based enzyme immunoassay (NSH-EIA), which integrates the heptamerization of antibody and enzyme. The Nb/SBP heptamer was generated by prokaryotic expression, during which the heptamerization was completed by self-assembly through the α-chains of C4-binding protein (C4bpα). Owing to the strong affinity of Nb/SBP heptamer and high enzyme load of SA-polyHRP, the NSH-EIA shows a limit of detection of 0.065 ng/mL with the linear range of 0.125–8 ng/mL. The NSH-EIA is highly selective for AFP and shows satisfactory accuracy and precision for the analysis of spiked serum samples. Moreover, the NSH-EIA was used to test clinical serum samples and confirmed by the automated chemiluminescence immunoassay analyzer with a good correlation. Thus, this study demonstrated that the Nb/SBP heptamer is a valuable immunoreagent and the NSH-EIA could be a useful diagnostic tool for ultrasensitive detection of AFP and other serum biomarkers.

The serological diversity of serum IgG/IgA/IgM against SARS-CoV-2 nucleoprotein, spike, and receptor-binding domain and neutralizing antibodies in patients with COVID-19 in Japan.

BackgroundWe compared the temporal changes of immunoglobulin M (IgM), IgG, and IgA antibodies against severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) nucleoprotein (N), spike 1 subunit (S1), and receptor‐binding domain (RBD), and neutralizing antibodies (NAbs) against SARS‐CoV‐2 in patients with coronavirus disease 2019 (COVID‐19) to understand the humoral immunity in COVID‐19 patients for developing drugs and vaccines for COVID‐19.MethodsA total of five confirmed COVID‐19 cases in Nissan Tamagawa Hospital in early August 2020 were recruited in this study. Using a fully automated chemiluminescence immunoassay analyzer, we measured the levels of IgG, IgA, and IgM against SARS‐CoV‐2 N, S1, and RBD and NAbs against SARS‐CoV‐2 in COVID‐19 patients' sera acquired multiple times in individuals from 0 to 76 days after symptom onset.ResultsIgG levels against SARS‐CoV‐2 structural proteins increased over time in all cases but IgM and IgA levels against SARS‐CoV‐2 showed different increasing trends among individuals in the early stage. In particular, we observed IgA increasing before IgG and IgM in some cases. The NAb levels were more than cut‐off value in 4/5 COVID‐19 patients some of whose antibodies against RBD did not exceed the cut‐off value in the early stage. Furthermore, NAb levels against SARS‐CoV‐2 increased and kept above cut‐off value more than around 70 days after symptom onset in all cases.ConclusionOur findings indicate COVID‐19 patients should be examined for IgG, IgA, and IgM against SARS‐CoV‐2 structural proteins and NAbs against SARS‐CoV‐2 to analyze the diversity of patients' immune mechanisms.

Association Between First-Trimester Maternal Cytomegalovirus Infection and Stillbirth: A Prospective Cohort Study.

BackgroundGiven that the time lag between cytomegalovirus (CMV) screening and diagnosed testing, a better knowledge of the association between pregnant women with CMV screening test positive and stillbirth in an epidemiological perspective was required to assist people being counseled reframe their pregnancy and birth plans based on the magnitude of the risk.MethodsThis study recruited 44048 eligible pregnant women from March 13, 2013 to December 31, 2019. Serological tests including CMV-specific IgM and IgG, and IgG avidity index were used to screen for maternal CMV infection and were measured by automated chemiluminescence immunoassay. The association was assessed using the inverse probability of group-weighted multivariate-adjusted log-binomial models.ResultsA total of 540 infants ended with a stillbirth (12.3 per 1000 pregnancies), and 2472 pregnancies with maternal CMV infection were screened out (56.1 per 1000 pregnancies) among all eligible pregnancies. In the comparison analysis, 326 infants ended with a stillbirth (86.6 per 1000 pregnancies) in the maternal CMV infection group compared with 214 infants (7.8 per 1000 pregnancies) in the group where mothers were not infected with CMV (RR 12.17; 95% CI 9.43–15.71). After excluding the pregnancies of stillbirth with birth defects, a strong association between the two groups was still observed (RR 9.38; 95% CI 6.92–12.70).ConclusionOur findings quantified the risk of a woman having a baby with stillbirth if she had a positive serologic CMV screening test in her first trimester, and supported the value of using CMV serologic tests as part of regular testing in pregnant women.Trial registrationRegistered in Chinese Clinical Trial Registry Center; registration number, ChiCTR1800016635; registration date, 06/14/2018 (Retrospectively registered); URL of trial registry record, https://www.chictr.org.cn/showproj.aspx?proj=28300.

Persistence of SARS-CoV-2 total immunoglobulins in a series of convalescent plasma and blood donors.

BackgroundThe vast majority of COVID-19 cases both symptomatic and asymptomatic develop immunity after COVID-19 contagion. Whether lasting differences exist between infection and vaccination boosted immunity is yet to be known. The aim of this study was to determine how long total anti-SARS-CoV2 antibodies due to past infection persist in peripheral blood and whether sex, age or haematological features can influence their lasting.Material and methodsA series of 2421 donations either of SARS-CoV-2 convalescent plasma or whole blood from 1107 repeat donors from January 2020 to March 2021 was analysed. An automated chemiluminescence immunoassay for total antibodies recognizing the nucleocapsid protein of SARS-CoV-2 in human serum and plasma was performed. Sex, age, blood group, blood cell counts and percentages and immunoglobulin concentrations were extracted from electronic recordings. Blood donation is allowed after a minimum of one-month post symptom’s relapse. Donors were 69.7% males and their average age was 46. The 250 donors who had later donations after a positive one underwent further analysis. Both qualitative (positivity) and quantitative (rise or decline of optical density regarding consecutive donations) outcomes were evaluated.Results and discussionIn 97.6% of donors with follow-up, anti-SARS-CoV-2 protein N total antibodies remained positive at the end of a follow-up period of 12.4 weeks median time (1–46, SD = 9.65) after the first positive determination. The blood group was not related to antibody waning. Lower lymphocyte counts and higher neutrophils would help predict future waning or decay of antibodies. Most recovered donors maintain their total anti-SARS-CoV-2 N protein antibodies for at least 16 weeks (at least one month must have been awaited from infection resolution to blood donation). The 10 individuals that could be followed up longer than 40 weeks (approximately 44 weeks after symptom’s relapse) were all still positive.

Generation of a nanobody-alkaline phosphatase heptamer fusion for ratiometric fluorescence immunodetection of trace alpha fetoprotein in serum

Alpha fetoprotein (AFP) is an important biomarker for diagnosis of hepatocellular carcinoma (HCC). Whereas, it is always a challenge to detect trace AFP in serum. In this work, a ratiometric fluorescence enzyme immunoassay (RFEIA) was developed using nanobody-alkaline phosphatase (Nb-AP) heptamer and MnFe layered double hydroxides nanoflakes (MnFe LDH) for ultrasensitive detection of AFP. The Nb-AP heptamer (Nb-C4bpα-AP) was constructed by fusion expression of Nb, AP, and α-chain of C4 binding protein (C4bpα), where the C4bpα contributed to multimerization through self-assembly. The dual functional Nb-C4bpα-AP can recognize AFP, dephosphorylate ascorbic acid-2-phosphate (AAP) into ascorbic acid (AA), and thus tune the MnFe LDH-mediated ratiometric fluorescence, which was generated from the oxidization of MnFe LDH on o-phenylenediamine (OPD) and the catalyzation of MnFe LDH on the cyclization reaction between AA and OPD. By integration of Nb-C4bpα-AP, MnFe LDH, AAP, and OPD, the RFEIA showed a limit of detection of 0.013 ng/mL with good selectivity, accuracy and precision. Furthermore, results of clinical serum samples tested by the RFEIA were well confirmed by the automated chemiluminescence immunoassay analyzer. Thus, this work demonstrated that the Nb-C4bpα-AP is a robust immunoreagent and the developed RFEIA could be a very promising tool for diagnosis of HCC.

Re-evaluation of the diagnostic value and optimal cutoff point of captopril challenge test in diagnosis of primary aldosteronism

Objective: The aim of the present study was to re-evaluate the diagnostic value and optimal cutoff point of captopril challenge test (CCT) in diagnosis of primary aldosteronism (PA). Methods: This is a retrospective study. All patients with a high risk for PA underwent screening test, and then proceeded to CCT and fludrocortisone suppression test (FST) on different days. The FST was used as a reference standard for PA. The plasma renin concentration (PRC) and plasma aldosterone concentration (PAC) were measured with an automated chemiluminescence immunoassay. Random number method was performed in the patients with unilateral primary aldosteronism (UPA), in order to make the proportion of the analyzed UPA in PA was 35%. Receiver operating characteristic (ROC) analyses were performed to compare diagnostic accuracy. Results: A total of 543 patients with 400 PA patients and 143 essential hypertension (EH) patients were enrolled. The diagnostic value of post-CCT PAC was significantly higher than that of the post-CCT plasma aldosterone-renin ratio (ARR), and that of the PAC suppression percentage, respectively. The area under the ROC curve (AUCROC) was 0.86 (0.83, 0.89) for PAC, 0.78 (0.74, 0.82) for ARR, and 0.62 (0.56, 0.67) for the PAC suppression percentage (all P<0.01), respectively. The optimal cutoff point of post-CCT PAC for PA was 110 ng/L, in which the sensitivity and specificity were 73.25% and 79.02%, respectively. The diagnostic efficiency of post-CCT PAC was not improved either in combination with PAC suppression percentage or in combination with post-CCT ARR. Conclusions: CCT is a useful test for the confirmation of PA. PAC level of 110 ng/L at 2 h after 50 mg of captopril is recommended as an optimal cutoff point for the diagnosis of PA.