IgG Titration Research Articles

In vitro data suggest that Indian delta variant B.1.617 of SARS-CoV-2 escapes neutralization by both receptor affinity and immune evasion.

BackgroundEmerged mutations can be attributed to increased transmissibility of the B.1.617 and B.1.36 Indian delta variants of SARS‐CoV‐2, most notably substitutions L452R/E484Q and N440K, respectively, which occur in the receptor‐binding domain (RBD) of the Spike (S) fusion glycoprotein.ObjectiveWe aimed to assess the effects of mutations L452R/E484Q and N440K (as well as the previously studied mutation E484K present in variants B.1.351 and P.1) on the affinity of RBD for ACE2, SARS‐CoV‐2 main receptor. We also aimed to assess the ability of antibodies induced by natural infection or by immunization with BNT162b2 mRNA vaccine to recognize the mutated versions of the RBD, as well as blocking the interaction RBD‐ACE2, an important surrogate readout for virus neutralization.MethodsTo this end, we produced recombinant wild‐type RBD, as well as RBD containing each of the mutations L452R/E484Q, N440K, or E484K (the latest present in variants of concern B.1.351 and P.1), as well as the ectodomain of ACE2. Using Biolayer Interferometry (BLI), we measured the binding affinity of RBD for ACE2 and the ability of sera from COVID‐19 convalescent donors or subjects immunized with BNT162b2 mRNA vaccine to block this interaction. Finally, we correlated these results with total anti‐RBD IgG titers measured from the same sera by direct ELISA.ResultsThe binding assays showed L452R/E484Q double‐mutant RBD to interact with ACE2 with higher affinity (KD = 4.6 nM) than wild‐type (KD = 21.3 nM) or single mutants N440K (KD = 9.9 nM) and E484K (KD = 19.7 nM) RBDs. Meanwhile, the anti‐RBD IgG titration resulted in lower recognition of mutants E484K and L452R/E484Q by infection‐induced antibodies, whereas only mutant E484K was recognized less by antibodies induced by vaccination. More interestingly, sera from convalescent as well as immunized subjects showed reduced ability to block the interaction between ACE2 and RBD mutants E484K and L452R/E484Q, as shown by the inhibition assays.ConclusionOur data suggest that the newly emerged SARS‐CoV‐2 variant B.1.617, as well as the better‐studied variants B.1.351 and P.1 (all containing a mutation at position E484) display increased transmissibility both due to their higher affinity for the cell receptor ACE2 and their ability to partially bypass immunity generated against the wild‐type virus. For variant B.1.36 (with a point mutation at position N440), only increased affinity seems to play a role.

Evaluation of survival rate using liposome containing soluble antigens (SA) against Toxoplasma gondii infection in BALB/c mice

Objective(s)Toxoplasma gondii, an obligate intracellular protozoan parasite, is widespread across the world. It causes congenital disease and abortion in humans and domestic animals. One of the major concerns in parasitology, thus, is an effective vaccine development to control Toxoplasmosis.Materials and MethodsIn the present research, a nano-liposomal vaccine containing soluble antigens (SA) was designed to evaluate the immunity and protective efficacy against T. gondii infection in BALB/c mice. Soluble antigens (SA) were achieved from tachyzoites, encapsulated in the liposome, and investigated via scanning electron microscope. Three times with 2-week intervals, BALB/c mice were immunized subcutaneously with different formulations. The level of protection against infection was assessed through the percent survival survey of BALB/c mice after challenge with tachyzoites of T. gondii RH strain; also, the type of generated immune response was determined by evaluating the generation of cytokine (IFN-γ, IL-4) and titration of IgG isotypes.ResultsThe immunization with liposome DSPC+ SA and liposome DSPC+ Imiquimod + SA induced a substantial increase in anti-Toxoplasma IgG antibody as compared to the PBS group (p <0.05). The IgG2a and IFN-γ secretion highest levels were seen with liposome DSPC+ Imiquimod + SA more than the control group (p <0.01) and (p <0.0001), respectively. After challenge with tachyzoites, less mortality was detected in the immunized mice by liposome DSPC + Imiquimod + SA that was meaningfully different (p <0.01) in comparison to other groups.ConclusionVaccination with liposome DSPC + Imiquimod + SA showed more survival rate and cellular immune reaction against T. gondii.

Immunogenicity of the Multi-Epitopic Recombinant Glycoproteins of Newcastle Disease Virus: Implications for the Serodiagnosis Applications.

BackgroundNewcastle disease virus (NDV) is a dangerous viral disease, infecting a broad range of birds, and has a fatal effect on the poultry industries. The attachment and consequently fusion of the virus to the host cell membrane is directed by the two superficial glycoproteins, the hemagglutinin-neuraminidase (HN) and the fusion (F) which is considered as the important targets for the poultry immune response.ObjectivesThe principal goal of this investigation was to realize the potential efficacy of the E. coli expression system for the production of the multi-epitopic HN, and F proteins with respect to the ability for the stimulation of the immune system and production of the cross-reactive antibodies in mice.Materials and MethodsThe recombinant HN and F (rHN, rF) have accumulated almost 40% of the total bacterial proteins. The presence of rHN and rF proteins recognized by the Western blotting with specific anti-HN, anti-F, anti-Newcastle B1, and anti-poly 6x His-tag antibodies. Furthermore, both rHN and rF have shown the specific reactivity against the Newcastle B1 antiserum as a standard strain.ResultsThe ELISA analysis showed that the higher dilutions of the antibody against Newcastle B1 could react with the as least quantity as 100 ng of the purified rHN, and rF. Cross-reactivity analysis of the sera from the mice immunized with Newcastle B1 in two time points indicated that the raise of anti-Newcastle B1, anti-HN and anti-F antibodies peaked at 28 days post immunization (dpi). Moreover, temporal variation in IgG titration between both time points was significant at 5% probability level.ConclusionThe results provided valuable information about the cross-reactivity patterns and biological activity of the multi-epitopic proteins compared to the NDV standard strain which was determined by the Western blotting and ELISA.

Rituximab-induced hypogammaglobulinemia in patients with neuromyelitis optica spectrum disorders

ObjectiveTo evaluate the long-term effects of rituximab (RTX) on total and specific immunoglobulins (Igs) in patients with neuromyelitis optica spectrum disorders (NMOSDs).MethodsTotal IgG, IgA, and IgM levels were evaluated in 15 patients with NMOSDs treated with RTX (median follow-up 70 months). Anti-aquaporin 4 (AQP4)-IgG titration was performed on samples from 9 positive patients. Anti-tetanus (TET), anti-varicella-zoster virus (VZV), and anti-Epstein–Barr virus nuclear antigen (EBNA) IgGs were also tested in patients with NMOSDs and in 6 healthy controls (HCs).ResultsRTX reduced total IgG by 0.42 g/L per year, IgA by 0.08 g/L per year, and IgM by 0.07 g/L per year. Hypogammaglobulinemia (hypo-IgG) (IgG < 7 g/L) developed in 11/15 patients. Severe hypo-IgG (IgG < 4 g/L) was found in 3/15 patients, of whom 2 patients developed serious infectious complications. In group analysis, anti-AQP4 IgG titers were reduced by RTX over time, and a significant correlation between anti-AQP4 IgG titers and total IgG levels was found. The effects of RTX were observed on pathogen-specific IgGs as well. In particular, the levels of anti-TET IgG in patients were significantly lower than those in HCs. The half-life of anti-TET IgG was reduced by about 50% in patients compared with the general population.ConclusionsLong-term RTX treatment is associated with the risk of hypo-Ig and reduction of anti-TET protection in patients with NMOSDs. Results obtained in this study suggest the importance of monitoring total and specific Ig levels before and during treatment with anti-CD20 drugs to prevent hypo-Ig–related complications and to optimize clinical management.

Serological diagnosis of mumps: Value of the titration of specific IgG

IntroductionThe aim of this study was to evaluate a cut-off point of the titration of IgG by ELISA in the diagnosis of mumps. MethodsA study was made of serum samples from 85 mumps cases (confirmed by PCR in saliva) and 2351 controls of the general population of the Region of Madrid. ResultsThe IgM detection was positive in 21 cases (sensitivity of 24.7%). The best cut-off point corresponded to IgG titres≥4900 (sensitivity of 64.7% and specificity of 86.1%). Among 42 patients immunised with at least one dose of measles mumps, rubella vaccine IgM was detected in 4 cases. However, the detection of IgG≥4900 was positive in 29 (sensitivity of 69.0%). ConclusionsAn IgG result of ≥4900 was almost 5 times more probable in a patient with mumps than in a non-infected patient. The detection of high titres of IgG against mumps could improve the IgM results in vaccinated people.

Diagnóstico serológico de parotiditis epidémica: valor de la titulación de IgG específica

IntroductionThe aim of this study was to evaluate a cut-off point of the titration of IgG by ELISA in the diagnosis of mumps. MethodsA study was made of serum samples from 85 mumps cases (confirmed by PCR in saliva) and 2,351 controls of the general population of the Region of Madrid. ResultsThe IgM detection was positive in 21 cases (sensitivity of 24.7%). The best cut-off point corresponded to IgG titres ≥4,900 (sensitivity of 64.7% and specificity of 86.1%). Among 42 patients immunised with at least one dose of measles mumps, rubella vaccine IgM was detected in 4 cases. However, the detection of IgG ≥4,900 was positive in 29 (sensitivity of 69.0%). ConclusionsAn IgG result of ≥4.900 was almost 5 times more probable in a patient with mumps than in a non-infected patient. The detection of high titres of IgG against mumps could improve the IgM results in vaccinated people.

Effects of oregano (Origanum vulgare L.) and rosemary (Rosmarinus officinalis L.) aqueous extracts on broiler performance, immune function and intestinal microbial population

ABSTRACTA 57-day study was performed to determine the effects of two aqueous extracts (AEs) on broiler performance, immune function and intestinal microflora. Four groups of 75 one-day-old female broilers (Ross308) received one of the following treatments: (1) a standard commercial feed (C); (2) C supplemented with 2 g/kg rosemary AE (R); (3) C supplemented with 2 g/kg oregano AE (O); (4) C supplemented with 1 g/kg oregano AE + 1 g/kg rosemary AE (OR). Individual body weight, average daily gain, average daily feed intake and feed conversion efficiency were determined at 1, 11, 22, 36 and 57 days. Sample collections for IgG titration and intestinal microflora examination were performed at 22 and 57 days. The addition of oregano AE alone or in combination with rosemary AEs improved body weight up to 36 days of age (P < .01). A time effect was recorded for total serum IgG in all groups (P < .001) and the percentage increase of the value was positively (P < .05) influenced by the AE supplementation. Lactobacilli raised (P < .001) in ileum and cecum of all groups supplemented with AEs. Staphylococcus spp. population was constantly lower in both intestinal tracts of the AE supplemented groups. On the basis of our results, AEs could improve broiler performance and immune function and contribute to a balanced gut microflora, essential for the digestion process and protection against enteropathogenic organisms.

Seroprevalence of Toxoplasmosis among Women Referring to Shahid Beheshti Hospital, Hamadan, Iran

Background: Toxoplasmosis is a parasitic disease caused by the protozoan Toxoplasma gondii. The aim of this study was investigate the prevalence of toxoplasmosis among young women who referred to check up for toxoplasmosis attended in Shahid Beheshti hospital, Hamadan during 2013-2014. M aterials and Methods: This study was performed on 2523 pregnant women who referred to laboratory of Shahid Beheshti hospital in Hamadan province (western of Iran) during 2013-2014. Age, level of education and place of residence were recorded in the relevant forms. Antibodies serum levels for all samples were examined by ELISA. IgG titer equals and more than 1:200 was presumed as seropositive. Data were analyzed using by SPSS version 19.0 software. R es ults: 26.1% of IgG seropositive persons were city residents while 32.3% of them lived at village and suburb of city. 1.4% and 1.1% of at risk persons (based on IgG titration) were city and village residents, respectively. 1.3% and 1.9% of IgM seropositives were city and village residents, respectively. The percentage of at risk persons of city and village (based on IgM titration) were 0.3% and 0.6%, in a row. 29.7% of IgG seropositives did not have academic education while 30.4% of them graduated from high school, at least. The seropositive IgM percentage of non-academic educated persons and graduated/academic ones were 1.7% and 1.4%, respectively. C onclusion: Our funding indicates the association between age of women and their level of education with percentage of contamination and prevalence. IgM seropositive is lesser than IgG. It means that toxoplasmosis is chronic or there is previous contact. To avoid the risk of toxoplasmosis infection particularly in pregnant women should be examined and the necessary preventive measures and training for young women should be presented.

Detection of Immunoglobulin G against E7 of Human Papillomavirus in Non-Small-Cell Lung Cancer

Background. A significant number of non-small-cell lung cancers (NSCLC) have human papillomavirus (HPV) DNA integrated in their genome. This study sought to further establish HPV's possible etiologic link to NSCLC by evaluating an immune response to HPV's oncogene, E7, in patients with NSCLC. Patients and Methods. Antibodies (IgG) in serum against E7 for HPV 16 and 18 in 100 patients with NSCLC were examined by enzyme-linked immunosorbent assay (ELISA). Results. Sixteen NSCLC patients were found to have a high titration of IgG for HPV oncogenic E7 protein. 23.5% of adenocarcinomas (AC,) and 15.4% of squamous cell carcinomas (SCC) were positive for IgG against HPV E7. HPV-18 (11%) had a slightly higher frequency than HPV-16 (6%). Of the six positive cases for HPV-16, 3 were AC, 2 SCC, and 1 NOS (not otherwise specified). For the 11 HPV-18 positives, 7 were AC, and 4 SCC. The one case with IgG against HPV 16 and 18 was AC. One case had high cross-reactive levels against E7 of HPV 16 and 18. Two (28%) of 7 patients who reported never smoking were positive for HPV, and 12 (13.6%) of 88 smokers were HPV positive. Conclusions. The study detected high levels of IgG against E7 in 16% of NSCLC patients. This adds evidence to a potential role of HPV in the pathogenesis of NSCLC.

Amamentação parcelada em função da ordem de parto de porcas

O objetivo foi avaliar os efeitos da amamentação parcelada nas primeiras seis horas após o nascimento, sobre o peso ao desmame, aos 60 dias de idade e a dosagem de imunoglobulinas G séricas em leitões de porcas de diversas ordens de parto. Trinta e três fêmeas suínas foram usadas em delineamento inteiramente casualizado e em esquema fatorial 2 × 3, composto de dois manejos de leitões pós-nascimento e três agrupamentos por ordem de parto. No manejo tradicional, os leitões disputavam as tetas e mamavam aleatoriamente, enquanto no manejo com alimentação parcelada foram distribuídos em dois grupos para mamar. A coleta de sangue dos leitões foi realizada seis horas após o término do parto para titulação de IgG. Aos 21 e 60 dias de idade, os leitões foram pesados. O manejo de alimentação parcelada não afetou o peso nem a dosagem de IgG sérica dos leitões, independentemente da ordem de parição das porcas, no entanto os leitões das porcas primíparas apresentaram menor peso ao desmame. O manejo de alimentação parcelada resultou em aumento da concentração de IgG sérica dos leitões de porcas primíparas. A amamentação parcelada deve ser utilizada em primíparas, mas não necessariamente em porcas de segunda parição em diante, pois os leitões de primíparas submetidos ao manejo de alimentação parcelada apresentam maiores concentrações de IgG sérica.

Sensitivity and specificity of immunoglobulin G titer for the diagnosis of mumps virus in infected patients depending on vaccination status

The aim of the study was to evaluate the usefulness of serological detection of mumps IgM and titration of IgG in patients with acute parotitis according to their vaccination status. The detection of mumps virus RNA in saliva by RT-PCR was used as reference. 116 patients (109 of them previously vaccinated) with mumps RT-PCR-negative results and 21 (19 vaccinated) with mumps RT-PCR-positive results were studied. Mumps-specific IgM and IgG were assayed by EIA (Enzygnost, Dade Behring, Germany). IgM results were expressed as positive or negative. For IgG, several cut-offs were calculated using receiver operating characteristic (ROC) curves. Seven RT-PCR-positive and five RT-PCR-negative patients showed IgM-positive results (sensitivity 33.3% and specificity 95.7%). Among vaccinated patients, the sensitivity and specificity of IgM were 26.3% (5/19) and 99.1% (108/109). For IgG, a titer of 5,000 in all the patients showed a sensitivity of 76.2% (16/21) and a specificity of 83.6% (97/116). In vaccinated patients, the corresponding figures for this cut-off were 84.2% (16/19) and 83.5% (91/109), respectively. Although IgM detection against mumps is highly specific, its sensitivity is very low in immunized subjects. In this group, the titration of IgG could serve as an additional diagnostic tool.

RSV molecular characterization and specific antibody response in young children with acute lower respiratory infection.

The presence of respiratory syncytial virus (RSV) in nasopharyngeal aspirates (NPA) were studied in 254 hospitalized Argentinean children with acute lower respiratory infection tract (ALRI). The specific humoral immune response and partial sequences of the G protein gene were studied in a subset of 22 children with RSV confirmed infection. The RSV IgM detection and the RSV IgG titration were made by immunofluorescence assay (IFA) in pairs of sera. The partial RSV G gene sequences were obtained by an RT-PCR amplification directly from de NPAs. RSV was present in 44.5% of the children. The RSV IgM was detected in 22.7 and 68.8% of the first and second sera, respectively. The IgG geometric mean titers of the acute and convalescent sera were 8 and 589. The RSV IgG titration was able to define 86.4% of the RSV confirmed cases. The percentage of coincidence between RSV IgM detection in the second sera and diagnosis by RSV IgG titration was 72.7% and no significant differences were observed. The nucleotide sequence of one group A and three group B viruses were identified. The first one was related with circulating viruses in Madrid, Montevideo and Mozambique during 1992, 1989 and 1999, respectively. The three sequences identified as group B viruses were closely related with circulating viruses in 1998 from South Africa and Canada during 1999 and 2000. The data obtained in our study provide the first approach at the molecular level (nucleotide) of the RSV circulating strains in Argentina and the lack of genotype patterns previously determined make necessary a continuous molecular surveillance in order to contribute to the understanding of the behavior of this virus in our community.

Parotiditis en un área urbana de la Comunidad de Madrid. Estado vacunal, diagnóstico y medidas de intervención

To describe the epidemiological situation of an epidemic of mumps, to calculate the risk of catching the illness depending on the vaccine strain used and to evaluate the diagnostic performance of the serologic determination. Longitudinal observational study. The incidence of mumps between January 1999 and June 2000 was calculated. Case-control study of outbreaks evaluated the risk of catching the disease depending on the vaccine strain. Health Area 4 in the Community of Madrid. Residents and students from schools in the district.Interventions. Epidemiological vigilance. Serologic determination of IgM and titration of IgG. In outbreaks, people exposed were vaccinated after assessment of their vaccine status. The rate of incidence in the first 27 weeks of the year 2000 for the 1-4 year-old age group was 334.7/105, against 12.2 in 1999. In children under 5 vaccinated with the Rubini strain there was a significantly greater risk of suffering illness than in those vaccinated with the Jeryl-Lynn strain (OR, 6.28, 95% CI, 1.43-38.50). The amount of cases confirmed serologically (specific IgM positive) was 22.22%. The mean of the logarithms of the IgG titres was significantly higher (p < 0.001) in IgM-negative patients with epidemiological confirmation (4.05, 95% CI, 3.18-4.28) than in IgM-negative cases without any epidemiological link (2.58; 95% CI, 2.33-2.84). We emphasise the increase of mumps incidence in the school population (fundamentally in the under-5s), which parallels an increase in outbreaks. The results suggest that the Rubini strain is less efficacious. The possibility of using alternative methods to IgM detection (IgG titration) as a diagnostic tool in a population with high vaccination coverage is posed.

Antitumor activity of specific immunotherapy with mucin containing CA 125 antigen in mice with CaO 1 ovarian carcinoma.

Experiments in CBA mice with transplanted CaO 1 ovarian carcinoma possessing common antigenic determinants with human ovarian carcinoma showed that specific immunotherapy with mucin containing CA 125 antigen inhibited tumor growth by 60% and prolonged animal lifespan by 40-60% in comparison with the control. The correlation coefficient between the tumor size and antibody titer after injection of mucin was -0.4 for IgM and -0.6 for IgG. Titration of IgG may be used for monitoring of the efficiency of specific immunotherapy.

An enzyme-linked immunosorbent assay (ELISA) for IgG and IgA antibodies to respiratory syncytial virus in low dilutions of secretions of human serum and secretions.

An enzyme-linked immunosorbent assay (ELISA) has been developed for titration of IgG and IgA antibodies to respiratory syncytial (RS) virus in low dilutions of human serum, colostrum, and nasopharyngeal secretions. Previously the sensitivity of RS virus ELISA on such specimens has been limited by nonspecific absorption of antibody, particularly IgA, to crude antigen preparations. For IgG antibody estimation in infant sera, this unwanted binding was reduced to workable levels by increasing the serum, salt, and detergent concentration of the diluent. Residual nonspecific binding of IgA in colostra appeared mainly due to antigen lipids or to lipoproteins. This was markedly reduced by partitioning Triton X-100-treated infected cell lysate antigens in Arklone. Using the modified ELISA technique for anti-RS virus IgA, good correlations were found with unfixed cell membrane immunofluorescence (MIF) for colostra (r = 0.81, P less than 0.001) and nasal secretions from adult volunteers. In several samples nonspecific absorption of antibody precluded MIF assay, but did not affect the ELISA. Although there was an overall correlation between ELISA for anti-RS IgG antibody in sera, the complement fixation test (r = 0.75, P less than 0.001), and MIF test (r = 0.82, P less than 0.001), the sensitivity of ELISA for antibody responses in convalescent sera of infants from 3 months to 2 years was poor. Conversely, the sensitivity of ELISA for antibody in the sera of older children and for transplacentally acquired antibody in very young infants was higher than that for the other two tests. ELISA was thus less reliable than either CF or MIF for detecting antibody rises in paired infant sera, particularly where maternally acquired antibody remained in the acute serum. The reasons for this apparent disparity are discussed.

An ELISA technique to detect IgG antibody to the early herpes simplex virus type 2 (HSV-2) antigen AG-4 in HSV-2 patients

An ELISA system, based on urease activity was used for the detection and titration of IgG to the immediate early AG-4 antigen in sera from HSV-2 patients. It detected low levels of IgG to the AG-4 antigen in 32% of patients'sera known to contain complement fixing antibody to the antigen. Furthermore, the sensitivity of the ELISA system was 2-to 10-fold higher than the complement-fixation system depending on when the sera was taken from the HSV-2 patients. The system also allowed the easy detection and quantitation of AG-4 antigen production when various HSV-1 X HSV-2 intertypic recombinant viruses were used to infect BHK-21 cells.

A quantitative determination for the detection of immunoglobulin (IgG) on the surface of platelets

A sensitive method for the quantitation of IgG on platelets had not been demonstrated until 1975, when Dixon, Rosse, and Ebbert described a quantitative antiglobulin consumption test useful in detecting platelet associated IgG (N Engl J Med 292:230, 1975). A modification of that technique has rendered the assay reproducible and removed the need for daily repetition of a standard IgG titration curve for quantitation. This modification utilizes 1-ethyl-3-3(dimethylaminopropyl)carbodiimide HCl (ECDI) (Sigma, E-7750), in place of chromic chloride, as a coupling agent for attaching IgG (Miles 64–145) to sheep cells (SRC), used as indicator cells. The ECDI consistently couples IgG to SRC and does not subject the SRC to sporadic spontaneous lysis, as does chromic chloride. This modification permits the detection of IgG on platelets (Direct Test), or in sera (Indirect Test) by incubation of a washed platelet pool with sera in vitro, and testing as in the Direct Test. Normal values of 0.01–1.56 and 0.14–1.6 femtograms (F) per platelet have been obtained for the Direct and Indirect Tests, respectively. In six cases of suspected ITP, values ranged 12.0–221.0 F and 2.9–37.6 F for the Direct and Indirect Tests, respectively. In conclusion, in disease states or other abnormal situations, quantities of IgG can be detected that are not usually present on the platelets of normal subjects.