Role of anti-polyethylene glycol (PEG) antibodies in the allergic reactions to PEG-containing Covid-19 vaccines: Evidence for immunogenicity of PEG

A small fraction of recipients who receive polyethylene-glycol (PEG)-containing COVID-19 mRNA-LNP vaccines (Comirnaty and Spikevax) develop hypersensitivity reactions (HSRs) or anaphylaxis. A causal role of anti-PEG antibodies (Abs) has been proposed, but not yet been proven in humans. We used ELISA for serial measurements of SARS-CoV-2 neutralizing Ab (anti-S) and anti-PEG IgG/IgM Ab levels before and after the first and subsequent booster vaccinations with mRNA-LNP vaccines in a total of 291 blood donors. The HSRs in 15 subjects were graded and correlated with anti-PEG IgG/IgM, just as the anti-S and anti-PEG Ab levels with each other. The impacts of gender, allergy, mastocytosis and use of cosmetics were also analyzed. Serial testing of two or more plasma samples showed substantial individual variation of anti-S Ab levels after repeated vaccinations, just as the levels of anti-PEG IgG and IgM, which were over baseline in 98–99 % of unvaccinated individuals. About 3-4 % of subjects in the strongly left-skewed distribution had 15–45-fold higher values than the median, referred to as anti-PEG Ab supercarriers. Both vaccines caused significant rises of anti-PEG IgG/IgM with >10-fold rises in about ∼10 % of Comirnaty, and all Spikevax recipients. The anti-PEG IgG and/or IgM levels in the 15 vaccine reactors (3 anaphylaxis) were significantly higher compared to nonreactors. Serial testing of plasma showed significant correlation between the booster injection-induced rises of anti-S and anti-PEG IgGs, suggesting coupled anti-S and anti-PEG immunogenicity. Conclusions: The small percentage of people who have extreme levels of anti-PEG Ab in their blood may be at increased risk for HSRs/anaphylaxis to PEGylated vaccines and other PEGylated injectables. This risk might be further increased by the anti-PEG immunogenicity of these vaccines. Screening for anti-PEG Ab “supercarriers” may help predicting reactors and thus preventing these adverse phenomena.     

Deciphering IgM interference in IgG anti-HLA antibody detection with flow beads assays

In flow beads assays, the interference of IgM for IgG anti-HLA antibodies detection is not precisely understood. Using the screening flow beads assay for class I HLA antibodies, we analyzed the binding of two IgG mAbs, the anti-class I HLA W6/32 and an anti-beta-2-microglobulin, in the presence of an anti-beta-2-microglobulin IgM mAb. In neat serum, the IgM mAb impaired the detection of both IgG. In EDTA-treated serum, the interference was stronger for the anti-beta-2-microglobulin IgG than for W6/32, in agreement with the finding in surface plasmon resonance that this IgM competed with the anti-beta-2-microglobulin IgG but not with W6/32. The IgM interference was higher in neat than in EDTA-treated serum for both IgG mAbs. The IgM interference was also analyzed with class II single antigen flow beads and sera from two kidney recipients containing IgG and IgM donor specific antibodies. Anti-HLA IgG detection was partially corrected by EDTA, and restored by IgM inactivation with DTT, confirming the results observed with the mAbs. Therefore, three mechanisms can explain the IgM interference for IgG anti-HLA antibodies in flow beads assays: direct competition for antigen, steric hindrance and complement activation.     

Preparation and evaluation of a recombinant Rift Valley fever virus N protein for the detection of IgG and IgM antibodies in humans and animals by indirect ELISA

This paper describes the cloning, sequencing and bacterial expression of the N protein of the Rift Valley fever virus (RVFV) ZIM688/78 isolate and its evaluation in indirect ELISAs (I-ELISA) for the detection of IgM and IgG antibodies in human and sheep sera. Sera used for the evaluation were from 106 laboratory workers immunised with an inactivated RVF vaccine, 16 RVF patients, 168 serial bleeds from 8 sheep experimentally infected with wild type RVFV and 210 serial bleeds from 10 sheep vaccinated with the live attenuated Smithburn RVFV strain. All human and animal sera that tested positive in the virus neutralisation test were also positive in the IgG I-ELISA. There was a high correlation (R² = 0.8571) between virus neutralising titres and IgG I-ELISA readings in human vaccinees. In experimentally infected sheep IgG antibodies were detected from day 4 to 5 post-infection onwards and IgM antibodies from day 3 to 4. The IgG I-ELISA was more sensitive than virus neutralisation and haemagglutination-inhibition tests in detecting the early immune response in experimentally infected sheep. The I-ELISAs demonstrated that the IgG and IgM response to the Smithburn vaccine strain was slower and the levels of antibodies induced markedly lower than to wild type RVFV infection.     

The immunological benefit of higher dose N-acetyl cysteine following mechanical ventilation in critically ill patients

Background

Sepsis complication is a major cause of death in multiple trauma critically ill patients. Defensin (cysteine rich anti-microbial peptides), as an important component of immune system, might play an important role in this process. There is also rising data on immunological effects of N-acetyl-cysteine (NAC), a commonly used anti-oxidant in oxidative stress conditions and glutathione (GSH) deficiencies. The aim of the present study was to evaluate the potential beneficial effects of NAC administration on multiple trauma patients with sepsis.

Methods

In a prospective, randomized controlled study, 44 multiple trauma critically ill patients who were mechanically ventilated and met the criteria of sepsis and admitted to the intensive care unit (ICU) were randomized into two groups . Control group received all standard ICU therapies and NAC group received intravenous NAC 3 gr every 6 hours for 72 hours in addition to standard therapies. Acute Physiology and Chronic Health Evaluation II (APACHE II) and Sequential Organ Failure Assessment (SOFA) scores, length of ICU stay, ICU mortality were recorded. Levels of serum Immunoglobulin M (IgM), Human β-Defensin 2 (HβD2) and GSH were assessed at baseline and 24, 72, 120 hours after intervention.

Results

During a period of 13-month screening, 44 patients underwent randomization but 5 patients had to be excluded. 21 patients in NAC group and 18 patients in control group completed the study. For both groups the length of ICU stay, SOFA score and systemic oxygenation were similar. Mortality rate (40% vs. 22% respectively, p = 0.209) and ventilator days (Mean ± SD 19.82 ± 19.55 days vs. 13.82 ± 11.89 days respectively, p = 0.266) were slightly higher for NAC group. IgM and GSH levels were similar between two groups (p = 0.325, 0.125 respectively), HβD2 levels were higher for NAC group (at day 3).

Conclusion

High dose of NAC administration not only did not improve patients’ outcome, but also raised the risk of inflammation and was associated with increased serum creatinine.     

Diagnostic Potential of a Luminex-Based Coronavirus Disease 2019 Suspension Immunoassay (COVID-19 SIA) for the Detection of Antibodies against SARS-CoV-2

Due to the current, rapidly increasing Coronavirus disease 2019 (COVID-19) pandemic, efficient and highly specific diagnostic methods are needed. The receptor-binding part of the spike (S) protein, S1, has been suggested to be highly virus-specific; it does not cross-react with antibodies against other coronaviruses. Three recombinant partial S proteins of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) expressed in mammalian or baculovirus-insect cells were evaluated as antigens in a Luminex-based suspension immunoassay (SIA). The best performing antigen (S1; amino acids 16-685) was selected and further evaluated by serum samples from 76 Swedish patients or convalescents with COVID-19 (previously PCR and/or serologically confirmed), 200 pre-COVID-19 individuals (180 blood donors and 20 infants), and 10 patients with acute Epstein-Barr virus infection. All 76 positive samples showed detectable antibodies to S1, while none of the 210 negative controls gave a false positive antibody reaction. We further compared the COVID-19 SIA with a commercially available enzyme immunoassay and a previously evaluated COVID-19 rapid antibody test. The results revealed an overall assay sensitivity of 100%, a specificity of 100% for both IgM and IgG, a quantitative ability at concentrations up to 25 BAU/mL, and a better performance as compared to the commercial assays, suggesting the COVID-19 SIA as a most valuable tool for efficient laboratory-based serology.     

Different Cross-Reactivities of IgM Responses in Dengue,Zika and Tick-Borne Encephalitis Virus Infections

Flaviviruses circulate worldwide and cause a number of medically relevant human diseases, such as dengue, Zika, yellow fever, and tick-borne encephalitis (TBE). Serology plays an important role in the diagnosis of flavivirus infections, but can be impeded by antigenic cross-reactivities among flaviviruses. Therefore, serological diagnosis of a recent infection can be insufficiently specific, especially in areas where flaviviruses co-circulate and/or vaccination coverage against certain flaviviruses is high. In this study, we developed a new IgM assay format, which is well suited for the specific diagnosis of TBE, Zika and dengue virus infections. In the case of TBE and Zika, the IgM response proved to be highly specific for the infecting virus. In contrast, primary dengue virus infections induced substantial amounts of cross-reactive IgM antibodies, which is most likely explained by structural peculiarities of dengue virus particles. Despite the presence of cross-reactive IgM, the standardized nature and the quantitative read-out of the assay even allowed the serotype-specific diagnosis of recent dengue virus infections in most instances.     

应用重组抗原(rSAG1)检测IgM抗体诊断弓形虫病的研究

目的以刚地弓形虫RH株重组主要表面抗原1作为检测抗原,建立检测弓形虫IgM抗体的rSAG1-ELISA.方法用Ni2 螯合柱纯化重组抗原rSAGI;以不同浓度的rSAG1的包被聚苯乙烯酶标条,阳性和阴性混合血清分别作1:100和1:200稀释,以辣根过氧化物酶标记的羊抗人IgM为二抗,采用正交试验确定rSAG1-ELISA的最佳检测条件;按其最佳检测条件分别对弓形虫IgM阳性和阴性混合血清重复测20次进行精确度的检测;采用抗体抑制试验检测其特异性;同时对用进口试剂盒筛得的35份弓形虫IgM阳性血清和57份弓形虫IgM阴性血清进行检测.结果 制备的rSAG1重组蛋白纯度在90%以上;rSAG1-ELISA的最佳检测条件为:rSAG1的包被浓度为2.5μg/m1,血清稀释度1:100,酶标记的羊抗人IgM 1:4 000稀释;用rSAG1-ELISA对混合弓形虫IgM阳性和阴性血清的重复检测表明:IgM阳性混合血清检测值的变异系数(CV值)为13.8%,IgM阴性混合血清的CV值为7.7%;灵敏度检测表明血清稀释度在1:50～1:200均可检出阳性;特异性试验的抑制率为62.0%;rSAG1-ELISA与进口试剂盒的总符合率为88.0%,其中阳性和阴性符合率分别为82.9%(29/35)和91.2%(52/57).结论 rSAG1-ELISA具有较好的灵敏度和特异性,与进口试剂盒相比有较高的符合率,表明该法对弓形虫病早期诊断具有较高的价值,可进一步推广应用.     

Kinetics of immunoglobulin and specific antibody responses of CBA mice infected with Trypanosoma rhodesiense

Summary Groups of CBA/CaJ and B-cell deficient CBA/N mice were infected with Trypanosoma rhodesiense EATRO 1886 strain. Survival, parasitaemia, serum Ig levels plus specific trypanosomal IgM and IgG antibodies were assayed and compared during infection. Whereas both strains of mice had similar parasitaemias during the first week of infection, CBA/N parasitaemias were lower than those observed in CBA/CaJ mice during the subsequent study period. Antibody responses, specific for T. rhodesiense antigens, peaked on day 10 after infection in CBA/CaJ mice, then rapidly declined. However, antibody responses in CBA/N mice remained elevated throughout the study. In addition, the kinetics of specific IgG and IgM varied in CBA/N mice: IgG antibody was detected on day 4, whereas specific IgM was detected on day 16. This unique relationship between the appearance of IgG and IgM antibody may explain the longer survival observed for B-cell deficient CBA/N mice infected with T. rhodesiense.