Analysis of human chorionic gonadotropin (hCG): Application of routine immunological methods for initial testing and confirmation analysis in doping control

Human chorionic gonadotropin (hCG) is dimeric glycoprotein produced by placenta in pregnancy and also in low levels by pituitary gland. The main clinical use for exogenous hCG‐administration is typically linked to infertility. The desired effect of hCG misuse in sport is due to the enhancement of testicular production of testosterone. Therefore, hCG is listed by the World Anti‐Doping Agency (WADA) as a prohibited substance in male athletes and according to the recently published WADA guideline urinary concentrations of hCG > 5 IU/L may be an indicator of doping. In this study two independent immunoassays were used to implement the new WADA guideline. The assay for initial testing (Siemens Immulite 2000 XPi hCG assay) recognizes various hCG variants (e.g. hCG and β‐core fragment of hCG) whereas the confirmatory assay (PerkinElmer DELFIA Xpress hCG) is sensitive to intact and nicked hCG only. Both assays showed adequate sensitivity and were proven fit‐for‐purpose in routine doping control. Population‐based distribution of the assays was in good agreement with results of earlier studies and supported well the current threshold of 5 IU/L. Copyright © 2013 John Wiley & Sons, Ltd.     

Diagnostic value of anti-GBV-C antibodies in HIV-infected patients

The beneficial effect of co-infection by GB virus C (GBV-C) in the course of the disease in human immunodeficiency virus (HIV)-infected patients has been described, although its mechanism of action is yet to be determined. The role of anti-GBV-C antibodies in HIV-infected patients also remains unknown. At present, there are no commercial systems to detect specific markers of GBV-C infection. The research presented follows our previous work from which we obtained chimeric molecules formed by two domains of different GBV-C proteins with good sensitivity/specificity balances in the detection of anti-GBV-C antibodies in hemodialyzed and chronic hepatitis patient samples. It has been investigated the ability of the synthetic peptides to recognize specific anti-GBV-C antibodies in HIV and HCV/HIV co-infected patients by a peptide-based ELISA immunoassay. The results showed that human immunodeficiency virus-infected patients have a significantly higher frequency of anti-GBV-C antibodies than healthy controls. A comparison between HCV(+) /HIV(+) and HCV(-) /HIV(+) was analyzed. Although a higher percentage of HCV/HIV-positive sera were positive for antibodies against GBV-C peptides, the difference was not significant. The presence of anti-GBV-C antibodies could represent a good marker of exposure to GBV-C in HIV-infected patients to facilitate a further analysis of the effect of this exposure in the progression of illness caused by HIV infection.     

Urokinase receptor forms in serum from non-small cell lung cancer patients: relation to prognosis

Objectives

To study the prognostic impact of the different forms of the receptor for urokinase plasminogen activator (uPAR) in serum from 171 non-small cell lung cancer (NSCLC) patients.

Materials and methods

Serum sampled preoperatively was available from 171 patients radically resected for NSCLC. Intact uPAR, uPAR(I-III), intact and cleaved uPAR, uPAR(I-III) + uPAR(II-III) and the liberated uPAR(I) were measured by time-resolved fluorescence immunoassays (TR-FIAs 1-3).

Results

High serum levels of each of the three uPAR forms were associated with short overall survival (OS). In a multivariate survival analysis uPAR(I-III) (hazard ratio (HR) = 2.3, 95% confidence interval (CI): 1.2-4.5, p = 0.015) and uPAR(I) (hazard ratio (HR) = 1.5, 95% CI: 1.0-2.2, p = 0.0497) remained significant prognostic parameters independent of stage, histology, age, performance status and therapy.

Conclusions

This retrospective study shows that uPAR(I-III) and uPAR(I) in serum are independent prognostic factors in patients radically operated for NSCLC. Further prospective studies are needed to validate these markers for clinical use.     

Performance evaluation of three automated quantitative immunoassays and their correlation with a surrogate virus neutralization test in coronavirus disease 19 patients and pre‐pandemic controls

BackgroundSARS‐CoV‐2 pandemic is currently ongoing, meanwhile vaccinations are rapidly underway in some countries. The quantitative immunoassays detecting antibodies against spike antigen of SARS‐CoV‐2 have been developed based on the findings that they have a better correlation with the neutralizing antibody.MethodsThe performances of the Abbott Architect SARS‐CoV‐2 IgG II Quant, DiaSorin LIAISON SARS‐CoV‐2 TrimericS IgG, and Roche Elecsys anti‐SARS‐CoV‐2 S were evaluated on 173 sera from 126 SARS‐CoV‐2 patients and 151 pre‐pandemic sera. Their correlations with GenScript cPass SARS‐CoV‐2 Neutralization Antibody Detection Kit were also analyzed on 173 sera from 126 SARS‐CoV‐2 patients.ResultsArchitect SARS‐CoV‐2 IgG II Quant and Elecsys anti‐SARS‐CoV‐2 S showed the highest overall sensitivity (96.0%), followed by LIAISON SARS‐CoV‐2 TrimericS IgG (93.6%). The specificities of Elecsys anti‐SARS‐CoV‐2 S and LIAISON SARS‐CoV‐2 TrimericS IgG were 100.0%, followed by Architect SARS‐CoV‐2 IgG II Quant (99.3%). Regarding the correlation with cPass neutralization antibody assay, LIAISON SARS‐CoV‐2 TrimericS IgG showed the best correlation (Spearman rho = 0.88), followed by Architect SARS‐CoV‐2 IgG II Quant and Elecsys anti‐SARS‐CoV‐2 S (all rho = 0.87).ConclusionsThe three automated quantitative immunoassays showed good diagnostic performance and strong correlations with neutralization antibodies. These assays will be useful in diagnostic assistance, evaluating the response to vaccination, and the assessment of herd immunity in the future.     

全自动快速微粒子酶免疫检测法在肝移植中的应用

目的 探讨全自动快速微粒子酶免疫检测法（ＭＥＩＡ）在肝移植中的应用。方法 采用ＭＥＩＡ对１３例肝移植患者术 后不同时期血清中巨细胞病毒（ＣＭＶ）ＩｇＧ、ＩｇＭ的动态进行观察。结果 ＣＭＶ感染率在肝移植患者中高达３０－６５％。结论 ＭＥＩＡ具有特异性强、灵敏度高、重复性好、快速简便等优点,为目前最新最理想的ＣＭＶＩｇＧ、ＩｇＭ检测方法。     

The molecular relationship between antigenic domains and epitopes on hCG

Antigenic domains are defined to contain a limited number of neighboring epitopes recognized by antibodies (Abs) but their molecular relationship remains rather elusive. We thoroughly analyzed the antigenic surface of the important pregnancy and tumor marker human chorionic gonadotropin (hCG), a cystine knot (ck) growth factor, and set antigenic domains and epitopes in molecular relationships to each other. Antigenic domains on hCG, its free hCGα and hCGβ subunits are dependent on appropriate inherent molecular features such as molecular accessibility and protrusion indices that determine bulging structures accessible to Abs. The banana-shaped intact hCG comprises ∼7500 Å² of antigenic surface with minimally five antigenic domains that encompass a continuum of overlapping non-linear composite epitopes, not taking into account the C-terminal peptide extension of hCGβ (hCGβCTP). Epitopes within an antigenic domain are defined by specific Abs, that bury nearly 1000 Å² of surface accessible area on the antigen and recognize a few up to 15 amino acid (aa) residues, whereby between 2 and 5 of these provide the essential binding energy. Variability in Ab binding modes to the contact aa residues are responsible for the variation in affinity and intra- and inter-species specificity, e.g. cross-reactions with luteinizing hormone (LH). Each genetically distinct fragment antigen binding (Fab) defines its own epitope. Consequently, recognition of the same epitope by different Abs is only possible in cases of genetically identical sequences of its binding sites. Due to combinatorial V(D)J gene segment variability of heavy and light chains, Abs defining numerous epitopes within an antigenic domain can be generated by different individuals and species. Far more than hundred Abs against the immuno-dominant antigenic domains of either subunit at both ends of the hCG-molecule, the tips of peptide loops one and three (Ł1 + 3) protruding from the central ck, encompassing hCGβŁ1 + 3 (aa 20–25 + 64 + 68–81) and hCGαŁ1 (aa 13–22; Pro16, Phe17, Phe18) plus hCGαŁ3 (Met71, Phe74), respectively, have been identified in the two “ISOBM Tissue Differentiation-7 Workshops on hCG and Related Molecules” and in other studies. These Abs recognize distinct but overlapping epitopes with slightly different specificity profiles and affinities. Heterodimeric-specific epitopes involve neighboring αŁ1 plus βŁ2 (hCGβ44/45 and 47/48). Diagnostically important Abs recognize the middle of the molecule, the ck (aa Arg10, Arg60 and possibly Gln89) and the linear hCGβCTP “tail” (aa 135–145; Asp139, Pro144, Gln145), respectively. Identification of antigenic domains and of specific epitopes is essential for harmonization of Abs in methods that are used for reliable and robust hCG measurements for the management of pregnancy, pregnancy-related disease and tumors.