Amounts Of IgG Research Articles

Relationship between HLA-II Gene Polymorphisms and Immune Response in COVID-19 Survivors and Volunteers Vaccinated against This Infection.

The polymorphism of HLA class II genes was studied in COVID-19 survivors and vaccinated people. Six allelic variants of DQA1, 10 of DQB1, and 11 of DRB1 were identified. The DQA1*05:01 allele predominated in vaccinated volunteers and the DQA1*02:01, *01:03, and DQB1*06:02-8 alleles predominated in convalescents. In the groups with DQA1*01:01, *05:01 and *06:01 alleles, a high proportion of individuals seropositive for SARS-CoV-2 S protein was found. In the group with DQA1*02:01 allele, 40% of volunteers were found to have IgM and a significantly lower quantity of IgG to N protein of coronavirus. Comparative analysis showed a statistically significant increase in IL-10 levels in carriers of the DQA1*01:01 allele, which may indicate the influence of this allele on cytokine production. Further study of the immune response indices and their association with HLA class II gene polymorphism will provide better understanding of the mechanisms of COVID-19 immunogenesis.

Application of Interferon-γ Release Assay in the Assessment of T-Cell Immunity to SARS-CoV-2 Antigens in the Cohort of Pediatric Patients with Juvenile Idiopathic Arthritis.

Background: an accurate assessment of the immunity against SARS-CoV-2 can facilitate a better understanding and management of not only the recent coronavirus but similar pathogens as well. Objective: the aim of this study was to evaluate T-cell immunity with reference to antibody titers in a group of pediatric patients with autoimmune arthritides utilizing the widely known Interferon-γ Release Assay (IGRA). Materials and Methods: This study was conducted in the cohort of 55 children suffering from Juvenile Idiopathic Arthritis (JIA). This research analyzed the SARS-CoV-2 T-cell response measured by a specific quantitative IGRA, followed by a serological ELISA test measuring the presence and quantity of IgG, IgM, and IgA antibodies in serum. Results: The cellular response to SARS-CoV-2 measured by the IGRA test significantly correlated with the antibody titers, IgA (p < 0.00003, R = 0.537), IgG (p < 0.0001, R = 0.668), and IgG nucleocapsid protein (NCP) (p < 0.003, R = 0.0399), with no correlation with IgM levels. The antibody levels in patients receiving biological agents were significantly lower compared to the rest of the cohort (p = 0.0369), while traditional disease-modifying antirheumatic drugs had no such effect. Limitations: the main limitation of the research is the small sample size, mostly due to the specific cohort of patients and the lack of a healthy control. Conclusions: IGRA appears to be a viable tool in the accurate evaluation of T-cell responses to SARS-CoV-2, and serodiagnostics alone is not always sufficient in the assessment of immune responses.

ZMapp reduces diffusion of Ebola viral particles in fresh human cervicovaginal mucus

ABSTRACT Vaginal transmission from semen of male Ebola virus (EBOV) survivors has been implicated as a potential origin of Ebola virus disease (EVD) outbreaks. While EBOV in semen must traverse cervicovaginal mucus (CVM) to reach target cells, the behaviour of EBOV in CVM is poorly understood. CVM contains substantial quantities of IgG, and arrays of IgG bound to a virion can develop multiple Fc-mucin bonds, immobilizing the IgG/virion complex in mucus. Here, we measured the real-time mobility of fluorescent Ebola virus-like-particles (VLP) in 50 CVM specimens from 17 women, with and without ZMapp, a cocktail of 3 monoclonal IgGs against EBOV. ZMapp-mediated effective trapping of Ebola VLPs in CVM from a subset of women across the menstrual cycle, primarily those with Lactobacillus crispatus dominant microbiota. Our work underscores the influence of the vaginal microbiome on IgG-mucin crosslinking against EBOV and identifies bottlenecks in the sexual transmission of EBOV.

The Impact of Pathogen Reduction on Total IgG and IgG Subclass Profiles of Convalescent Plasma

Introduction: In case of newly emerging pathogens, convalescent plasma (CP) is often the only early available treatment option. It has been shown that different IgG subclasses contribute differently to CP neutralizing activity. As CP donors often have a risk profile like first-time donors, especially with respect to window-period viral transmission, pathogen reduction (PR) could mitigate that risk. The aim of our study, especially in the light of potential future pandemics, was to evaluate the impact of commercially available PR technologies on total IgG and IgG subclasses quantity and distribution in CP using COVID-19 CP (CCP) as surrogate for CP in a side-by-side comparison approach. Methods: 36 apheresis CCP donations were allocated to three study groups and a side-by-side assessment of the potential impact of amotosalen (AS)/UVA treatment compared to a riboflavin (RB)/UVB treatment, AS against methylene blue (MB) treatment, and RB against MB treatment on the quantity of IgG and IgG subclasses with a nephelometric analyzer was performed. Results: IgG subclass distributions were not significantly changed post PR treatment with all three technologies. There was also no significant difference in the median loss of concentration for IgG1 and IgG2 between the three technologies. We recognized a non-significant trend of a higher IgG4 median loss post RB treatment compared to post AS and MB treatment, respectively. Conclusion: Although the three commercially available PR systems do not significantly alter the distribution of IgG subclasses, we detected a non-significant trend of higher IgG4 loss after RB treatment. The potential impact of that finding needs further investigation.

Effects of enriching IgG concentration in low- and medium-quality colostrum with colostrum replacer on IgG absorption in newborn Holstein calves

Ingestion and absorption of greater quantities of IgG are required to increase serum IgG levels in newborn calves. This could be achieved by adding colostrum replacer (CR) to maternal colostrum (MC). The objective of this study was to investigate whether low and high-quality MC can be enriched with bovine dried CR to achieve adequate serum IgG levels. Male Holstein calves (n = 80; 16/treatment) with birth body weights (BW) of 40 to 52 kg were randomly enrolled to be fed 3.8 L of the following combinations: 30 g/L IgG MC (C1), 60 g/L IgG MC (C2), 90 g/L IgG MC (C3), C1 enriched with 551 g of CR (60 g/L; 30-60CR), or C2 enriched with 620 g of CR (90 g/L: 60-90CR). A subset of 40 calves (8/treatment) had a jugular catheter placed and were fed colostrum containing acetaminophen at a dose of 150 mg/kg of metabolic body weight, to estimate abomasal emptying rate per hour (kABh). Baseline blood samples were taken (0 h), followed by sequential samples at 1, 2, 3, 4, 5, 6, 8, 10, 12, 24, 36, and 48 h relative to initial colostrum feeding. Results for all measurements are presented in the following order, unless otherwise stated: C1, C2, C3, 30-60CR, and 60-90CR. Serum IgG levels at 24 h were different among calves fed C1, C2, C3, 30-60CR, and 60-90CR: 11.8, 24.3, 35.7, 19.9, and 26.9 mg/mL ± 1.02 (mean ± SEM), respectively. Serum IgG at 24 h increased when enriching C1 to 30-60CR, but not from C2 to 60-90CR. Similarly, apparent efficiency of absorption (AEA) values for calves fed C1, C2, C3, 30-60CR, and 60-90CR were different: 42.4, 45.1, 43.2, 36.3, and 33.4% ± 1.93, respectively. Enriching C2 to 60-90CR reduced AEA, and enriching C1 to 30-60CR tended to decrease AEA. The kABh values for C1, C2, C3, 30-60CR, and 60-90CR were also different: 0.16, 0.13, 0.11, 0.09, and 0.09 ± 0.005, respectively. Enriching C1 to 30-60CR or C2 to 60-90CR reduced kABh. However, 30-60CR and 60-90CR have similar kABh compared with a reference colostrum meal (90 g/L IgG, C3). Even though kABh was reduced for 30-60CR, results indicate that C1 has the potential to be enriched and achieve acceptable serum IgG levels at 24 h without affecting AEA.

Immunoassay for quantification of antigen-specific IgG fucosylation.

SummaryBackgroundImmunoglobulin G (IgG) antibodies serve a crucial immuno-protective function mediated by IgG Fc receptors (FcγR). Absence of fucose on the highly conserved N-linked glycan in the IgG Fc domain strongly enhances IgG binding and activation of myeloid and natural killer (NK) cell FcγRs. Although afucosylated IgG can provide increased protection (malaria and HIV), it also boosts immunopathologies in alloimmune diseases, COVID-19 and dengue fever. Quantifying IgG fucosylation currently requires sophisticated methods such as liquid chromatography-mass spectrometry (LC-MS) and extensive analytical skills reserved to highly specialized laboratories.MethodsHere, we introduce the Fucose-sensitive Enzyme-linked immunosorbent assay (ELISA) for Antigen-Specific IgG (FEASI), an immunoassay capable of simultaneously quantitating and qualitatively determining IgG responses. FEASI is a two-tier immunoassay; the first assay is used to quantify antigen-specific IgG (IgG ELISA), while the second gives FcγRIIIa binding-dependent readout which is highly sensitive to both the IgG quantity and the IgG Fc fucosylation (FcγR-IgG ELISA).FindingsIgG Fc fucosylation levels, independently determined by LC-MS and FEASI, in COVID-19 responses to the spike (S) antigen, correlated very strongly by simple linear regression (R2=0.93, p < 0.0001). The FEASI method was then used to quantify IgG levels and fucosylation in COVID-19 convalescent plasma which was independently validated by LC-MS.InterpretationFEASI can be reliably implemented to measure relative and absolute IgG Fc fucosylation and quantify binding of antigen-specific IgG to FcγR in a high-throughput manner accessible to all diagnostic and research laboratories.FundingThis work was funded by the Stichting Sanquin Bloedvoorziening (PPOC 19-08 and SQI00041) and ZonMW 10430 01 201 0021.

Abstract 514: Specific anti-HER2 host immunity conferred by HER2-targeted antibody drug conjugate therapy and checkpoint blockade

Abstract Using HER2-targeted antibody drug conjugates (ADC), we tested the therapeutic efficacy, direct impact on cancer stem cells (CSCs), and potential added benefit when combined with PD-L1 blockade in immunocompetent mouse tumor models. Two murine mammary tumor models were used in this study: D2F2/E2, a cell line resulting from stable transfection of parental D2F2 murine mammary tumor cells with wild-type human HER2, and 4T1-HER2, derived from parental 4T1 by stable transduction with wild-type human HER2. We demonstrate that HER2-targeted ADC treatment induced both cellular and humoral anti-HER2 adaptive immune responses which effectively targeted both bulk tumor and cancer stem cells. We recently investigated induction of host immunity using a tumor re-challenge strategy in tumor free animals subjected to ADC or ADC + anti-PD-L1 therapy. D2F2/E2 tumor-bearing mice were treated with ADC or ADC + anti-PD-L1 to generate tumor free mice. These tumor free animals were re-challenged with the relevant tumor D2F2/E2 or irrelevant tumor WT 4T1 respectively. While all the tumor free animals re-challenged with WT 4T1 showed aggressive tumor growth, ADC-treated tumor free animals re-challenged with D2F2/E2 demonstrated significantly (p&amp;lt;0.0001) slower tumor growth compared with the age-matched naive mice, and this protection was significantly (p=0.0078) enhanced in ADC + anti-PD-L1-treated tumor free animals, resulting in completely rejection of the re-challenge and thus documenting immunological memory formation. B cell activation was obvious by the significant upregulation of Ki67 and GL7 CD86 on both CD45+CD19+ tumor-infiltration lymphocytes (TILs) and splenocytes. B cell differentiation was evident by the upregulation of GL7 on both TILs and splenocytes, as well as the expression of CD138 on splenotytes. We found that IgG1 was the predominant isotype in treated mice vs. IgG2a, IgG2b, IgG3, and ADC + anti-PD-L1 dual treatment drastically increased the amount of IgG1 in serum. In addition, ADC+ anti-PD-L1 dual treatment significantly increased the frequency of CD8+IFNγ+ cells in TILs. Both ADC alone (p=0.0004) and ADC+anti-PD-L1 dual treatment (p=0.01) significantly decreased CD4+FoxP3+ regulatory T cells. These results provide evidence that HER2-targeted ADC + anti-PD-L1 immunotherapy-induced B cell activation and differentiation along with the modulation of T cell subsets could confer host anti-HER2 immunity. Citation Format: You Qin, Hannah E. Dobson, Frank I. Comer, Alfred E. Chang, Max S. Wicha, Qiao Li. Specific anti-HER2 host immunity conferred by HER2-targeted antibody drug conjugate therapy and checkpoint blockade [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 514.

Low pH Exposure During Immunoglobulin G Purification Methods Results in Aggregates That Avidly Bind Fcγ Receptors: Implications for Measuring Fc Dependent Antibody Functions.

Evaluating the biophysical and functional nature of IgG is key to defining correlates of protection in infectious disease, and autoimmunity research cohorts, as well as vaccine efficacy trials. These studies often require small quantities of IgG to be purified from plasma for downstream analysis with high throughput immunoaffinity formats which elute IgG at low-pH, such as Protein G and Protein A. Herein we sought to compare Protein G purification of IgG with an immunoaffinity method which elutes at physiological pH (Melon Gel). Critical factors impacting Fc functionality with the potential to significantly influence FcγR binding, such as IgG subclass distribution, N-glycosylation, aggregation, and IgG conformational changes were investigated and compared. We observed that transient exposure of IgG to the low-pH elution buffer, used during the Protein G purification process, artificially enhanced recognition of Fcγ Receptors (FcγRs) as demonstrated by Surface Plasmon Resonance (SPR), FcγR dimer ELISA, and a functional cell-based assay. Furthermore, low-pH exposed IgG caused conformational changes resulting in increased aggregation and hydrophobicity; factors likely to contribute to the observed enhanced interaction with FcγRs. These results highlight that methods employed to purify IgG can significantly alter FcγR-binding behavior and biological activity and suggest that the IgG purification approach selected may be a previously overlooked factor contributing to the poor reproducibility across current assays employed to evaluate Fc-mediated antibody effector functions.

An investigation into the factors associated with ewe colostrum production

The majority of lamb mortality which occurs during the first 24 h post-partum is preventable through providing the lamb with sufficient quantities of high quality colostrum during this time. Data from seven late gestation nutrition experiments carried out at this institute between 2002 and 2014 were collated into a single data set comprising of 415 twin bearing ewes. Analysis was carried out to investigate the key drivers of ewe colostrum production excluding nutrient intake, namely body reserve mobilisation, ewe breed type, ewe age, gestation length and lamb birth weight. The volume of colostrum produced at 1 and 18 h post-partum was significantly lower than the volume recorded at 10 h post-partum (P = 0.01). Multivariate regression analysis indicated that colostrum volume during the first 18 h post-partum was influenced by lamb birth weight (P = 0.01), ewe age (P = 0.01), breed type (P = 0. 01) and gestation length (P = 0.06). Live weight change (P = 0.05) also had a significant influence on the volume of colostrum produced but BCS change did not affect colostrum production (P = 0.25). Further multivariate regression analysis indicated that IgG yield was influenced ewe breed type (P = 0.01), lamb birth weight (P = 0.02), gestation length (P = 0.05) and BCS change (P = 0.04). Live weight change (P = 0.12) and ewe age (P = 0.62) did not influence the quantity of IgG produced. Leicester ewes produced less colostrum per kg lamb birth weight at 1 h post-partum compared to all other ewe breed types (P = 0.01) and less than Suffolk ewes at 10 h post-partum (P = 0.01). The result of this analysis shows the key factors excluding ewe nutrition that drive colostrum production. Ewe breed type in particular appears to play an important role in the ability of the ewe to produce sufficient quantities of adequate quality colostrum. In conclusion the result of this analysis highlights the important factors associated with ewe colostrum volume and IgG yield excluding nutrition. In particular the overall structure of the flock such as breed type and ewe age is important when considering the ability of the flock to meet colostrum demands and hence reduce lamb mortality.

Antigenic Epitope Analysis and Efficacy Evaluation of GRA41 DNA Vaccine Against T. gondii Infection.

Toxoplasma gondii has a comprehensive impact on a great range of warm-blood mammals, in which one-third of the population all over the world is involved. Dense granular proteins, regarded as GRA family, mediating substantial interface between host cell cytoplasm and parasite, are widely studied for preventing the infection of T. gondii. As is handled in our study, the effect of intramuscularly injecting the genetic vaccine pEGFP-C1/GRA41 encoding a novel dense granule protein, GRA41, was evaluated. At the beginning, bioinformatics analysis was used to evaluate epitopes of both B cells and T cells on the GRA41 protein of T. gondii. Afterwards, recombinant plasmids (pEGFP-C1/GRA41) were injected into BALB/c mice and the quantity of IgG and its subclass IgG2a remarkably increased. IFN-γ, distinctive from the other cytokines (IL-4, and IL-10), was significant in growth. Afterwards, the intraperitoneal challenge was executed for recording survival time with tachyzoites with high virulence (in RH strain) and counting the number of brain cysts was carried out after the infection of PRU strain (low virulence). In pEGFP-C1/GRA41 group, the survival period was significantly longer (13.3 ± 3.37 days) after tachyzoites attack with the RH strain in high virulence, compared with the other groups (less than 8 days). Additionally, the cyst quantity is remarkably lower and the rate of reduction could reach 59.34%. All the results indicated effective protection of DNA vaccine encoding GRA41 against T. gondii.

Optimization of Enzymatic Antibody Fragmentation for Yield, Efficiency, and Binding Affinity.

Enzymatic antibody fragmentation has been well studied for various hosts and isotypes, but fragmentation patterns also vary unpredictably by clone, and optimizing Fab or F(ab')2 production by trial and error consumes large quantities of antibodies. Here, we report a systematic strategy for optimizing functional F(ab')2 production via pepsin digestion from small quantities of IgG. We tested three key parameters that affect fragmentation, pH, enzyme concentration (% pepsin w/w), and reaction time, and found that pH had the greatest impact on fragmentation yield and efficiency. We then developed a systematic approach to obtaining acceptable yields, digestion efficiency, and binding affinity. Three case studies are described to illustrate the approach. We anticipate that this work will provide a quick and cost-effective method for researchers to produce antibody fragments from whole IgG, avoiding haphazard trial and error.

Concept of sample-specific correction of immunoassay results for precise and accurate IgG quantification in horse plasma

The hyperimmune horse plasma (HHP), prepared through active immunisation of horses with an antigen of interest, is the most common starting material for antitoxin (animal antibody-based therapeutics) production. Precise IgG quantification in plasma is a prerequisite for accurate estimation of the purification process efficiency. Although immunoglobulins from HHP have been purified for over a century, there is still no in vitro method for precise and accurate determination of IgG content in HHP. For this reason, the purification process efficiency has been assessed by antibody activity measurements, mostly performed in vivo. Here we describe the development of a precise and accurate in vitro immunoassay for IgG quantification in HHP. We showed and highlighted that any difference in composition of IgG population between the standard and the sample, with respect to both IgG subclass distribution and antigen-specific IgG content, leads to inaccurate IgG quantification. We demonstrated that caprylic acid precipitation as the method for IgG isolation from horse plasma renders the composition of IgG population unchanged. This very efficient, fast, simple and inexpensive method was used to prepare internal, sample-specific reference IgG for each plasma sample, which was tested simultaneously to a respective plasma sample. Deviation of IgG quantity determined by ELISA for each sample-specific reference from its nominal value was used for correction of the results of respective plasma sample, which led to accurate and precise IgG quantification as shown by method validation. The here presented novel concept of sample-specific correction of immunoassay results could be widely applicable and easily introduced in different immunoassays for more accurate and precise plasma IgG quantification.

The pattern of IgG concentration in neonatal lamb's sera from birth till 7th day (168th hour)

A quantitation of IgG was made, using R.I.D test, in blood sera of 15 newborn iambs. The quantity of IgG was measured in time intervals immediately after birth and before suckling till 168th hour after birth. Two lambs out of 15 have detectable amount of IgG in their sera before suckling. The maximum concentration of IgG in 11 lambs sera was measured 24 hours, in 3 lambs sera 36 hours, and in 1 lamb's serum 48 hours after birth, respectively. In three lambs out of 15, a percentage of 20%, the serum concentration of IgG was less than 30 mg/ml. One of these lambs died and another got ill 120 hours after birth.

An ELISA for the determination of human IgG based on the formation of a colored iron(II) complex and photometric or visual read-out

The paper describes a colorimetric sandwich enzyme-linked immunosorbent assay (ELISA) for the determination of human IgG. It is based on the use of an Fe(II) coordination complex as a signal amplifier and of mesoporous silica nanoparticles modified with glucose oxidase (GOx) and secondary antibodies (Ab2). After formation of the immuno sandwich complex, the quantity of GOx is proportional to the quantity of IgG. On addition of Fe(II) and glucose, GOx catalyzes the oxidation of glucose to produce hydrogen peroxide which oxidizes Fe(II) to Fe(III). After adding a stop solution containing the complexing ligand 1,10-phenanthroline (Phen), un-reacted Fe(II) forms an orange-red complex with Phen which can be detected by plate reader and even seen with bare eyes. This sandwich ELISA has a linear response in the 1 pg.mL−1 to 100 ng.mL−1 human IgG concentration range and a 860 fg.mL−1 detection limit. This is 20 times lower than the commercial ELISA for human IgG. The assay also is selective, stable, highly sensitive and cost-effective.

Serum bactericidal assay for the evaluation of typhoid vaccine using a semi-automated colony-counting method

Typhoid fever, mainly caused by Salmonella enterica serovar Typhi (S. Typhi), is a life-threatening disease, mostly in developing countries. Enzyme-linked immunosorbent assay (ELISA) is widely used to quantify antibodies against S. Typhi in serum but does not provide information about functional antibody titers. Although the serum bactericidal assay (SBA) using an agar plate is often used to measure functional antibody titers against various bacterial pathogens in clinical specimens, it has rarely been used for typhoid vaccines because it is time-consuming and labor-intensive. In the present study, we established an improved SBA against S. Typhi using a semi-automated colony-counting system with a square agar plate harboring 24 samples. The semi-automated SBA efficiently measured bactericidal titers of sera from individuals immunized with S. Typhi Vi polysaccharide vaccines. The assay specifically responded to S. Typhi Ty2 but not to other irrelevant enteric bacteria including Vibrio cholerae and Shigella flexneri. Baby rabbit complement was more appropriate source for the SBA against S. Typhi than complements from adult rabbit, guinea pig, and human. We also examined the correlation between SBA and ELISA for measuring antibody responses against S. Typhi using pre- and post-vaccination sera from 18 human volunteers. The SBA titer showed a good correlation with anti-Vi IgG quantity in the serum as determined by Spearman correlation coefficient of 0.737 (P < 0.001). Taken together, the semi-automated SBA might be efficient, accurate, sensitive, and specific enough to measure functional antibody titers against S. Typhi in sera from human subjects immunized with typhoid vaccines.

PO-17 - Identification of IgG bound to plasminogen in oncologic diseases

The binding of plasminogen (Pg) to cell receptors and extracellular ligands facilitates its activation to plasmin, which stimulates the extracellular matrix degradation, neoangiogenesis and tumor invasion. Plasmin can also degrade IgG thereby exposing C-terminal lysine residues. Previously, we have found IgG specifically bounded to Pg in the plasma of patients with malignant tumors. To identify IgG degraded by plasmin in the plasma of cancer patients. Methods of ELISA were used for comparative research of levels of IgG bound to Pg in plasma of patients with the prostate cancer (PC, n=25) and lung cancer (LC, n=17). Plasma of healthy donors (n=29) was used as control. All patients signed informed consent for participation in this study. Affinity chromatography on Pg-sepharose was used for the quantification of IgG. Carboxypeptidase was used for remove of C-terminal lysine residues of the IgG. The program ATTESTAT was used for nonparametric analysis. The frequency of occurence of elevated levels of IgG to Pg in plasma was detected in 68% of patients with PC, 59% of patients with LC and only 12% of healthy women and 10% of healthy men. The quantification of antibodies in plasma samples showed that the quantity of IgG to Pg in patients with PC was 27% from the total amount of IgG and in healthy men - 9%. Treatment of diluted plasma samples with carboxypeptidase B abolished the elevated levels of IgG to Pg, as well as the specific activity of the purified IgG to Pg-sepharose. C-terminal lysine residues which are formed as a result of degradation of native IgG with plasmin can bind to lysine binding sites on the kringle domains of Pg. Increased levels of these degraded IgG can be marker at cancer.

The discovery of IgE 50 years later

In 1906, Von Pirquet1 used the term supersensitivity without immunity to describe (1) symptoms of inhalant allergy, (2) positive immediate skin test results, and (3) the fact that the results of other tests of immunity were not positive in these patients. The other tests he referred to were precipitin tests and complement fixation. Thus, he opened up the question of serum tests for supersensitivity. In 1919, Ramirez2 reported that a patient who had received a blood transfusion from a horse allergic donor became allergic to horse dander. At this point, Prausnitz and Kustner3 set out to investigate whether the serum of allergic individuals contained a factor or factors that could sensitize the skin. In 1921, they reported that the local injection of serum from a fish allergic individual, Kustner, to an individual who was only allergic to pollen, Prausnitz, would transfer specific sensitivity.3 This transfer of sensitivity came to be known as the P-K test and was used widely to study sensitivity not only to common allergens but also to extracts as diverse as those obtained from schistosomes.4,5 Furthermore, in New York, Cooke et al6 identified that there were other antibodies in the serum that increased during desensitization treatment and could block the skin-sensitizing activity. By the 1950s, it was clear that the transferred sensitivity was specific, that it could be diluted extensively, and that the skin remained locally sensitive for days if not weeks after the injection of serum.7,8 It was also already clear that the ability to sensitize the skin was lost after moderate heating of the serum.7 Several studies had also been reported on the physical properties of P-K activity. Indeed, Heimlich et al9 at Cal Tech reported on the sedimentation properties of skin-sensitizing antibodies in 1960. However, the studies before 1964 had not succeeded in defining the nature of these antibodies.

Quantification of autoantibodies to plasminogen in plasma of patients with cancer.

The investigation of autoantibodies which may play a role in the processes of angiogenesis and tumorogenesis is important in the early diagnostis of cancer. This study aimed to investigate the levels of autoantibodies to Glu-plasminogen (Pg) in plasma of patients with tumors. Plasma samples from healthy volunteers were compared with samples from patients with prostate cancer using 2D electrophoresis and MALDI-TOF mass spectrometry. Plasma samples from 25 patients with prostate cancer, 15 patients with benign prostatic hyperplasia (BPH), 29 patients with breast cancer, and 43 healthy volunteers were tested using ELISA to anti-Pg IgG autoantibodies. Affinity chromatography on Pg-sepharoses was used to assess the quantity of anti-Pg IgG in control plasma and plasma of prostate cancer patients. ATTESTAT program was used for nonparametric analysis. Using 2D electrophoresis, marker spots below 50 kD were detected in prostate cancer samples. These spots were identified as fragments of Pg and IgG. Using affinity chromatography on Pg-sepharose, the quantity of IgG bound to Pg versus total IgG was determined to be 9% in control and 27% in prostate cancer samples. The frequency of occurence of elevated levels of anti-Pg IgG was 84% in prostate cancer samples, 69% in breast cancer samples, 40% in BPH samples, and 11% in healthy plasma. Autoantibodies to Pg may be involved in tumorogenesis and elevated levels of anti-Pg IgG antibodies may be a risk factor for tumor development.

Evaluation of a protocol to reduce the incidence of neonatal calf diarrhoea on dairy herds

Calf diarrhoea causes substantial economic losses in cattle herds worldwide. Neonatal calves are particularly sensitive to infections with enteropathogens. The present study focused on prevention against the main infectious causes of neonatal calf diarrhoea i.e. Escherichia coli, rota- and coronavirus, and Cryptosporidium parvum. Dairy herds (n=24) with a high percentage of neonatal calves scouring (>10%) were included and calves were sampled for the presence of these four enteropathogens. To decrease diarrhoea problems among neonatal calves, a standard protocol was tested on 13 herds (treatment group) where both C. parvum and either E. coli or rota- or coronavirus were identified as being involved, the other 11 herds served as control group. The protocol consisted of 2 points of action: preventive vaccination of dams against E. coli, rota- and coronavirus, and preventive administration of halofuginone lactate to newborn calves. The average percentage of calves suffering from neonatal diarrhoea (39.7% versus 14.3%, P<0.01) and the average percentage of faecal samples positive for C. parvum (34% versus 11%, P<0.05) differed significantly between control herds and treatment herds after implementation of the protocol. No significant differences between control and treatment group were observed in the percentage of calves excreting E. coli, rotavirus and coronavirus, both before and at the end of the trial. Furthermore, risk factors potentially associated with the development of neonatal calf scours were determined. Non-significant results were obtained for the effect of the protocol on duration of diarrhoea and the effect of the colostral IgG quantity on the risk of diarrhoea. Passive immunity transfer status of the calves, measured both before the onset and at the end of the study, were non-significant between groups.

AGE-DEPENDENT FEATURES OF EVOLVING HUMORAL IMMUNITY IN CHILDREN

Age dynamics of humoral immunity was studied in healthy children, i.e., 11 newborns, 33 infants of 4 to 8 months, 32 children of 1 to 2 years old,, 17 children of 4 to 5 years old, 25 children of 6 to 8 years old, 15 children of 9 to 11 years old, and 28 adolescents of 14 to 16 years old. Evaluation of membrane receptors on B cells was performed by means of three-colour fluorescent label and allowed of characterizing B1 subpopulations (CD19+CD5+CD27-), naпve B2 cells (CD19+CD5-CD27-), and B2 memory cells (CD19+CD5-CD27+). B1 cells have been shown to dominate in blood of newborns and younger children (up to 5 years old). By the contrary, B2 memory cells were nearly undetectable in newborns, and exceeded 20% in adolescents (by 15 years old). Meanwhile, it has been revealed that the amounts of IgG1 and IgG3 subclasses did progressively increase with age, whereas IgG2 remained decreased to 50% of adult values for a long time, and reached them by 11 years and later. We suggest that the age dynamics of IgG subclasses is connected with age-dependent changes in B cell subpopulations.