Time Course Of Antibody Response Research Articles

Characterization of antibody response in asymptomatic and symptomatic SARS-CoV-2 infection

BackgroundSARS-CoV-2 pandemic is causing high morbidity and mortality burden worldwide. Mechanism of infection, protection or rapid evolution until fatal outcome of the disease remains poorly understood. To elucidate the dynamics of humoral response in SARS-CoV-2 asymptomatic and symptomatic infections, we performed a study to investigate the time course of antibody response in hospitalized COVID-19 patients and asymptomatic SARS-CoV-2 infected subjects.MethodsSerum samples were collected from 42 hospitalized COVID-19 patients at 6 different time points (hospital admission, day 2, day 6, day 12-14, day 18-20, day 27-30) and from 25 asymptomatic subjects at 3 different time points (at the time of positive swab, 2 months, 6 months). Samples were tested by commercial ELISA for SARS-CoV-2 spike (S) protein IgA, IgM, and IgG and nucleoprotein (NP) IgG, and by micro-neutralization assay.ResultsIn patients, titres increased for all antibody classes including neutralizing antibody from day 6 to day 18-20 but at day 27-30 started to decline. A high correlation between S and NP antigens and among antibody classes was found. No significant difference in antibody titres at baseline and by peak antibody level was found between recovered and deceased. 64.0% of asymptomatic subjects were negative to any antibody at any time point. Asymptomatic subjects with positive antibody level had titres well below patients, and neutralizing antibodies were found only in 2 subjects (8.0%).ConclusionsOur results highlight that COVID-19 patients produce an antibody response to SARS-CoV-2 regardless the outcome. The peak is reached by 3 weeks from hospital admission followed by a sharp decrease. On the contrary, only few asymptomatic subjects develop antibodies at detectable levels, though lower compared to COVID-19 patients. Since neutralizing antibodies were rarely produced, this finding raises the question about the protection of these subjects against reinfection.Key messages COVID-19 patients produce an antibody response to SARS-CoV-2 which does not differ between recovered and deceased.Only few asymptomatic SARS-CoV-2 infected subjects develop antibodies and neutralizing antibodies were rarely produced.

Antibody response of definitive hosts against antigens of two life stages of the neuropathogenic schistosome Trichobilharzia regenti.

BackgroundThe nasal avian schistosome Trichobilharzia regenti spends part of its intravertebrate period of life within the central nervous system. Migration of the parasites can be accompanied by neuromotor disorders or paralysis in natural definitive hosts (ducks) and even in laboratory mammals. Cercariae are also able to penetrate human skin and induce cercarial dermatitis. While the cellular and antibody responses against cercariae and migrating schistosomula have been investigated in mice, little is known about immune reactions in birds. This study first describes the dynamics of antibody response in infected ducks and identifies frequently recognized antigens that may serve as diagnostic markers of infection by T. regenti.MethodsGroups of 35 domestic ducks and 10 mallards were exposed to different doses of T. regenti cercariae. Sera were collected at predefined time intervals and tested by ELISA for the presence of specific anti-cercarial IgY and IgM. Antigens recognized by the antibodies were identified on Western blots of cercariae and schistosomula. The applicability in immunodiagnostics was statistically evaluated by expression of specificity and sensitivity values for individual antigens.ResultsIn ELISA, the levels of anti-cercarial IgM peaked on day 15 pi. Increased production of IgY associated with the later phases of infection was observed in most individuals around 20 dpi and culminated 30 dpi. The time course of antibody response did not differ among experimental groups, variations were only observed in the levels of specific IgY which depended rather on the age of ducks at the time of infection than on the infectious dose. On Western blots, 40 cercarial and 7 schistosomular antigens were recognized by IgY from infected ducks. Among them, 4 cercarial antigens of 50, 47, 32 and 19 kDa provided the most sensitive and specific reactions.ConclusionsAntigens of cercariae and schistosomula elicited distinct antibody response in ducks, which correlated positively with the age of animals at the time of infection. Several antigens originating in cercariae and fewer in schistosomula were recognized by IgY with diverse sensitivity and specificity; only a few seemed to be common to both stages. Four of them were considered as the most promising candidates for immunodiagnostics.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-015-1007-y) contains supplementary material, which is available to authorized users.

Evaluation of an in-house TgSAG1 (P30) IgG ELISA for diagnosis of naturally acquired Toxoplasma gondii infection in pigs

Toxoplasma gondii is an apicomplexan protozoan parasite which is able to infect a large variety of warm-blooded animals. Raw or undercooked pork has been regarded as an important source of infection for humans. The aim of this study was to evaluate an in-house enzyme-linked immunosorbent assay to diagnose natural T. gondii infection in swine using native affinity chromatography-purified T. gondii surface protein-1 (TgSAG1-ELISA) as antigen, comparing its performance to that of indirect fluorescent antibody test (IFAT) and immunoblotting (IB). To obtain a panel of sera showing the evolution of the antibody response in the time course 12 pigs were experimentally inoculated intravenously (iv) with tachyzoites of the T. gondii strains RH (clonal type I), ME49 (clonal type II) and NED (clonal type III) and serologically monitored for a period of 11 weeks. Both IFAT and ELISA showed a similar time course of antibody response to T. gondii; but by IFAT this response was characterized by rapidly rising titers with peaks at two weeks post inoculation (wpi), while the ELISA indices increased slowly and reached a maximum in most animals at five wpi. Three-hundred randomly selected sera from a total of 602 pigs of different ages derived from outdoor and indoor farms from Argentina were analyzed. Serum samples testing either positive or negative by both IFAT and IB were considered as “relative standards of comparison” (RSC). Sensitivity and specificity of TgSAG1-ELISA were obtained by a Receiver Operating Characteristics (ROC) analysis and statistical agreement among serological tests was evaluated. Antibodies to T. gondii were detected in 160 of 300 sera (53.3%) by IB, in 133 of 300 (44.3%) by IFAT and in 123 of 300 sera (41%) by TgSAG1-ELISA. One hundred and eleven sera tested positive and 118 sera tested negative by both IFAT and IB (RSC); 103 of 111 positive RSC sera tested positive by TgSAG1-ELISA, and 116 of 118 negative RSC sera tested negative by TgSAG1-ELISA. Agreement observed between RSC and TgSAG1-ELISA was almost perfect (κ=0.9124, p≥0.05) and between IFAT and IB was moderate (κ=0.53, p≥0.05). Relative sensitivity and specificity of the TgSAG1-ELISA using a cut-off index of 0.204 were of 92.8% and 98.3%, respectively. ROC analysis revealed that TgSAG1-ELISA was highly accurate (AUC=0.983) relative to the RSC. According to the results in this study, the ELISA based on affinity purified T. gondii surface antigen TgSAG1 was useful for the specific and sensitive detection of antibodies to this protozoan parasite in naturally infected pigs.

Time course of antibody response to recombinant Aspergillus fumigatus antigens in cystic fibrosis with and without ABPA

We determined follow-up levels of specific serum IgE to the recombinant Aspergillus fumigatus (A. fumigatus) allergens rAsp f 1, 3, 4 and 6 in patients suffering from cystic fibrosis (CF) with and without allergic bronchopulmonary aspergillosis (ABPA). Over a 32-month period follow-up data of 74 patients were collected. According to serology, 11 CF patients were not sensitized (CF controls), 40 were sensitized to A. fumigatus (Asp. f-sens.) and 23 patients fulfilled the serologic criteria for ABPA. Of these 23 ABPA patients 11 expressed the full clinical ABPA picture (classicABPA) and 12 failed to show sufficient relevant clinical signs (seroABPA), despite positive serology. The 23 ABPA patients had 16-18 times higher serum levels of specific IgE to rAsp f 4 and/or rAsp f 6 than those of Asp. f-sens. patients (rAsp f 4: 31.3 +/- 45 EU/ml vs. 1.9 +/- 2.2 EU/ml and rAsp f 6: 39.0 +/- 44.3 EU/ml vs 2.1 +/- 1.7 EU/ml). The combination of increased total serum IgE (>1000 IU/l) and increased specific IgE to rAsp f 4 and/or rAsp f 6 allowed to diagnose classicABPA with 100% specificity and 64% sensitivity and with a high predicted positive (100%) and a high predicted negative (94%) value. During a combined treatment (seven patients) with oral corticosteroid and itraconazole, itraconazole alone (two patients) or neither oral corticosteroid nor itraconazole therapy (two patients) total serum IgE and specific IgE to rAsp f 4 and/or rAsp f 6 did decrease but did not normalize. Over the observation period, lung function remained unchanged, independent of whether oral steroids and/or concomitant itraconazole were either given or not given. In the follow-up of CF patients with ABPA under therapy the determination of total or specific IgE serum levels were of limited value to guide therapy.

Partial characterization of antigenic sperm proteins in foxes (Vulpes vulpes).

The aim of this work was to identify antigenic proteins on fox spermatozoa. Fox spermatozoa proteins were injected into 3 female rabbits and into 3 male and 3 female foxes. In rabbits, a rapid humoral response was observed. Using rabbit sera for Western blotting, 23 fox sperm protein bands were recognized between 10 and 110 kd. In foxes, the time course of antibody response was studied in the same manner. The number of recognized bands was maximal on day 75 for 2 foxes, on day 90 for 3 foxes, and on day 120 for 1 fox. Western blot patterns varied from one fox to another. On the whole, 25 protein bands between 10 and 110 kd were recognized. Using fluorescein isothiocyanate (FITC) labeling on fox spermatozoa with rabbit and fox sera, we showed that several antigens recognized by the antisera were located at or near the surface of the spermatozoa. By two-dimensional electrophoresis and gel-purification, we have selected 6 highly antigenic proteins with molecular weights of 11.4, 14.7, 16.4, 16.4, 16.8, and 16.9 kd, and isoelectric points of 6.0, 6.0, 6.2, 5.5, 5.3, and 5.8, respectively, and one antigenic protein at 97 kd with an isoelectric point of 4.3 to 4.6. The results of this study can be used to characterize these 7 antigens selected more precisely by microsequencing or mass spectrometry.

Identification of post-vasectomy sperm auto-antigens in fox ( Vulpes vulpes) by two-dimensional gel electrophoresis and Western blotting

The aim of this work was to identify antigenic surface proteins on fox spermatozoa. Six foxes were inguitinally vasectomised, and the time course of antibody response in the sera was studied. Five out of the six foxes reacted to vasectomy with a production of antisperm antibodies. The number of bands recognised by Western blot was maximal 120–150 days after the vasectomy, at the end of the reproductive season. On the whole, 30 bands were recognised between 9 and 150 kDa. The pattern of recognised proteins varied from one fox to another. The humoral response was studied in one fox 2 years after the vasectomy, before, during and after the breeding season. The same proteins were recognised, but the intensity of staining was increased during the testis regression. Using FITC-labelling on sperm smears with fox sera, antigens were localised at or near the sperm surface, either on the acrosome or both on the acrosome and on the flagellum. Using two-dimensional gel electrophoresis and Western blotting, we identified eight areas containing major antigens, recognised by at least two sera. The molecular weights (kDa) and isoelectric points of the repeated antigens were, respectively [150, 6.6–6.0]; [105–98, 6.0–5.5]; [97, 4.6–4.3]; [95, 5.0]; [85–80, 5.4–5.1]; [42, 5.0–4.8]; [17–15, 6.5–5.9]; and [17–15, 5.5–4.8]. The results of this study can be used to characterise more precisely fox sperm auto-antigens by microsequencing the selected proteins.

Time course of antibody response to tetanus toxoid and pneumococcal capsular polysaccharides in patients infected with HIV.

The temporal course of the humoral immune response to T-cell-dependent and T-cell-independent type 2 antigens was evaluated in HIV-infected patients. In all, 26 seropositive patients were vaccinated with tetanus toxoid and 23-valent pneumococcal vaccines; total IgG and IgG1 antibodies to tetanus toxoid (Ttox) and total IgG and IgG2 antibodies against 23 Streptococcus pneumoniae capsular antigens (PPS) were measured at baseline, 2 months, and 12 months after vaccination. For the Ttox, baseline levels of IgG1 (Ttox-IgG1) increased from 11.0 to 19.5 mg/L at 2 months postimmunization. Overall only 6 patients (23%) showed a significant response. At 12 months postvaccination, Ttox-IgG and T-tox-IgG1 were significantly lower than baseline levels (Ttox IgG basal; 11.0 mg/L, 12 months; 0.8 mg/L, Ttox IgG1 baseline; 13.1 mg/L, Ttox IgG1 12 months; 2.4 mg/L) and in 10 patients, antibodies that fell below protective levels (0.6 mg/L). In contrast with PPS, a significant response was observed at 2 and 12 months (PPS-IgG basal; 35.9 U/ml, 2 months; 151.4 U/ml, 12 months; 59.7 U/ml; PPS-IgG2 baseline 20.3 U/ml, 2 months; 113.2 U/ml, 12 months; 51.9 U/ml). Overall, 19 patients (76%) showed an immune response to pneumococcal polysaccharides antigens. Immunization with the Ttox T-cell-dependent antigen fails to elicit a significant immune response and may induce inhibition of antibody production in HIV-infected patients. In contrast, immunization with a T-cell-independent type 2 antigen can cause the pneumococcal polysaccharides to induce significant immune response in a high proportion of HIV-infected patients.

Acanthocheilonema viteae: vaccination with irradiated L3 induces resistance in three species of rodents (Meriones unguiculatus, Mastomys coucha, Mesocricetus auratus).

Three species of rodents were immunized with 50 irradiated (35 krad) stage-3 larvae (L3) of the filaria Acanthocheilonema viteae and challenged with an infection of normal L3. The immunization induced a significant reduction of the worm burden developing from the challenge infection in all host species, the jird (Meriones unguiculatus), the multimammate rat (Mastomys coucha) and the golden hamster (Mesocricetus auratus). The induced resistance was highest in jirds (92.5 +/- 9.7) followed by golden hamsters (59.4 +/- 26.6) and multimammate rats (55.1 +/- 40.4). The time course of antibody response against antigens of L3, adult worms and microfilariae, as studied by ELISA, showed quantitative and qualitative differences between the species. The antibody response against L3 antigens in immunoblots was similar in all species. Only one of the golden hamsters developed an antibody response against the surface of vector derived L3, while sera of jirds and multimammate rats did not react with L3 surface.

Antisperm autoantibody responses to vasectomy and vasovasostomy in Fischer and Lewis rats

Antisperm autoanitbodies were studied in Fischer and Lewis strains of rats after either vasectomy, vasectomy followed one month later by vasovasostomy, or sham operations. The time course of antibody response to sperm protein autoantigens was assayed by Western blot analysis of sera obtained at intervals up to 3 months. Rats of both strains responded to immunization with isologous spermatozoa with production of high titer hyperimmune sera. Sera from vasectomized Fischer rats showed antisperm antibodies on Western blots, but bands were stained with less intensity and frequency than for Lewis rats. In both Fischer and Lewis strains, major protein autoantigens were observed at 75–83, 68–71, 63, 57, 51, 41, and 21–23 kDa, lending support to the hypothesis that there is a set of dominant sperm autoantigens recognized by a consensus of postvasectomy rat sera. The lesser response of Fisher rats to vasectomy was not due to absence of dominant postvasectomy sperm autoantigens in Fischer sperm extracts, nor was it attributable to inability of Fischer rats to mount an immune response to these antigens, since immunization with isologous sperm was successful in raising antibodies to the dominant autoantigens. Vasovasostomy did not result in a general decrease in antisperm antibodies, and reactions to some antigens actually increased.

Measurement of isotype-specific antibody responses to Aujeszky's disease virus in sera and mucosal secretions of pigs

Enzyme-linked immunosorbent assays (ELISAs) for the detection of porcine IgM, IgA, IgG1 and IgG2 antibodies directed against Aujeszky's disease virus (ADV) are described. ADV-specific IgA and IgM were detected in an antibody capture assay, and ADV-specific IgG1 and IgG2 were detected in an indirect double antibody sandwich assay. A selected set of samples was tested in the four ELISAs and in a 24 h virus neutralization assay. Comparison of the results showed that the ELISAs were isotype-specific, sensitive, and reproducible. Samples with ADV antibody of one isotype showed that ADV-specific IgG1, IgG2 and IgM were able to neutralize the virus in vitro. In vitro neutralization of virus can be enhanced by complement. ADV-specific IgA neutralized virus only weakly. ADV-infected cells activated complement in the absence of antibody. Specific IgG2 and IgM enhanced complement activation. Analysis of the time course of antibody responses after infection or vaccination revealed that the isotype-specific ELISAs are suitable to study the humoral antibody response of pigs to the virus in mucosal secretions. Wild-type virus (strain NIA-3) and an attenuated vaccine strain (Bartha) administered intranasally induced mucosal IgM and IgA responses to the virus. In contrast, a killed vaccine (Nobivac) administered intramuscularly induced only weak mucosal IgM responses. The attenuated vaccine strain primed for a mucosal IgA memory response evoked upon challenge infection with wild-type virus.

Humoral immune responses to tumor-specific antigens in strain-2 guinea pigs.

Serum antibody against tumor-specific antigens on the surface of line-10 guinea-pig hepatoma cells was demonstrated by equilibrating serum with viable tumor cells and detecting bound antibody by indirect immunofluorescence. Summation of fluorescence intensity grades of individual cells led to a semiquantitative assay of antibody activity sufficiently precise for studies on the magnitude and time course of antibody response. Antibody was found not only in guinea pigs immunized against line-10 tumor with a line-10 BCG vaccine, but also in animals with progressively growing and metastasizing tumors. All animals studied had a primary humoral response to line-10 BCG vaccine and generally developed higher antibody levels after multiple inoculations of tumor cells. Immunization with line-10 antigen extracts in complete Freund adjuvant produced delayed cutaneous hypersensitivity responses, but no humoral antibody was detected until after challenge with viable tumor cells, when an apparent anamnestic antibody response was seen. Guinea pigs with progressively growing line-10 tumors developed antibody after 4-5 weeks of tumor growth. Before antibody was detected, their serum inhibited the binding of antitumor antibody to line-10 cells.