Specific Immunoglobulin Classes Research Articles

Full-Length Immune Repertoire Reconstruction and Profiling at the Transcriptome Level Using Long-Read Sequencing.

Due to the diversity of the immune repertoire (IR), reconstructing full-length IR using traditional short-read sequencing has proven challenging. A full-length IR sequencing (FLIRseq) work flow was developed with linear rolling circle amplification and nanopore sequencing. Its accuracy and quantification ability were verified by plasmid mixtures and commercial B-cell receptor/T-cell receptor sequencing (BCR/TCR-seq) based on short reads. IRs in tissues and the peripheral blood from 8 patients with acute lymphoblastic leukemia, 3 patients with allergic diseases, 4 patients with psoriasis, and 5 patients with prostate cancer were analyzed using FLIRseq. FLIRseq reads had lower mismatch rates and gap rates, and higher identify rates than nanopore reads (all P < 2.2 × -16). The relative quantification of components by FLIRseq was consistent with the actual quantification (P > 0.05). FLIRseq had superiority over BCR/TCR-seq, providing the long complementarity-determining region 3, B-cell isotype, and the rarely used V gene sequence. FLIRseq observed an increase in clonotype diversity (P < 0.05) and a decrease in the percentage of abnormal BCRs/TCRs in patients with leukemia in remission. For patients with allergic diseases or psoriasis, FLIRseq provided direct insights into V(D)J recombination and specific immunoglobulin classes. Compared with that in prostate cancer tissues, the full-length V segment of the biased T-cell receptor β chain from lymphocytes in psoriatic tissues showed a more consistent AlphaFold2-predicted protein structure (P < 0.05). FLIRseq enables unbiased and comprehensive analyses of direct V(D)J recombination and immunoglobulin classes, thereby contributing to characterizing pathogenic mechanisms, monitoring minimal residual disease, and customizing adoptive cell therapy.

Organization for laboratory diagnosis of SARS-COV-2 in the conditions of an anti-tuberculosis institution

Introduction. In connection with the spread of a new coronavirus infection COVID-19 since 2020, measures have been taken in the city of Moscow to widely diagnose the causative agent of COVID-19. A laboratory PCR department was opened in the State Budgetary Healthcare Institution «MNPC for the fight against tuberculosis of the DZM», taking into account the available material and human resources. Purpose of the study: To generalize the experience of organizing the laboratory service of an anti-tuberculosis institution in the city of Moscow during the spread of a new coronavirus infection. Results. Human and material resources have made it possible to maintain the quality and availability of laboratory diagnostics of biological material from patients receiving treatment at the Moscow Regional Research and Practical Center for Combating Tuberculosis, DZM, during the COVID-19 pandemic. In the period 2020-2022 in total, 90,102 studies of biological material (smears) were carried out by RT-PCR in order to detect COVID-19 RNA. Starting from June 2020 to the present, the PCR department of the KDL Clinic 2 has been studying the direct detection of Covid-19 RNA in biological material using molecular genetic methods based on reverse transcription and polymerase chain reaction (RT-PCR) and the identification of specific immunoglobulin classes IgM and IgG to coronavirus antigens in blood serum (plasma) by the method of immunochemiluminescent analysis (ICLA) of both persons receiving treatment at the State Budgetary Institution of Healthcare Center for Combating Tuberculosis Moscow, and employees.

Antibody responses to BNT162b2 SARS-CoV-2 mRNA vaccine among healthcare workers and residents of long-term care facilities: A cohort study in Northern Italy.

Long-term care facilities (LTCFs) have been severely impacted by COVID-19, with a disproportionate amount of SARS-CoV-2 infections and related deaths occurring among residents. This study is part of an ongoing multicenter, prospective cohort study conducted among healthcare workers(HCWs) and residents of 13 LTCFs in Northern Italy designed to evaluate SARS-CoV-2 specific immunoglobulin class G (IgG) titers before and following vaccination with Pfizer/BNT162b2 SARS-CoV-2 mRNA vaccine (two doses of vaccine, 21 days apart). Serum samples were obtained from participants (t0) before vaccination, and (t1) 2 weeks after and analyzed to determine anti-S1 IgG antibodies. Five hundred and thirty-fourparticipants were enrolled (404 subjects participated in both blood draws). Seropositivity was 50.19% at t0and 99% at t1, with a significant difference in IgG titers. A higher proportion of residents were seropositive at t0 compared withHCWs, with significantly higher IgG titers among residents at both t0 and t1. Pre-existing immunity also had a significant effect on postvaccination IgG titers. However, a significant difference in titers at t1 between HCWsand residents considering only participants seropositive at t0 was found, with higher median titers among previously seropositive residents. Findings of this study provide scientific evidence endorsing the policy of universal vaccination in LTCFs.

Affinity chromatography on monolithic supports for simultaneous and high-throughput isolation of immunoglobulins from human serum.

Posttranslational modifications of immunoglobulins have been a topic of great interest and have been repeatedly reported as a major factor in disease pathology. Cost-effective, reproducible, and high-throughput (HTP) isolation of immunoglobulins from human serum is vital for studying the changes in protein structure and the following understanding of disease development. Although there are many methods for the isolation of specific immunoglobulin classes, only a few of them are applicable for isolation of all subtypes and variants. Here, we present the development of a scheme for fast and simultaneous affinity purification of α (A), γ (G), and μ (M) immunoglobulins from human serum through affinity monolith chromatography. Affinity-based monolithic columns with immobilized protein A, G, or L were used for antibody isolation. Monolithic stationary phases have a high surface accessibility of binding sites, large flow-through channels, and can be operated at high flow rates, making them the ideal supports for HTP isolation of biopolymers. The presented method can be used for HTP screening of human serum in order to simultaneously isolate all three above-mentioned immunoglobulins and determine their concentration and changes in their glycosylation pattern as potential prognostic and diagnostic disease biomarkers.

Serological and virological evaluation of human T-lymphotropic virus type 1 infection in family groups from Tumaco, Colombia.

To date there has been no statistical evaluation of the profiles of immunoglobulin classes and viral replication as variables in the study of HTLV-1 infection and circulation among families in virus-endemic areas of Colombia. To evaluate the correlation of several immunological and molecular characteristics with the transmission and circulation of HTLV-1 among families in the town of Tumaco. Plasma levels of HTLV-1 specific immunoglobulin classes IgG, IgM and IgA1, as well as IgG and sIgA in oral fluids, were calculated for 32 members of 10 family groups from Tumaco in which the mother and at least one child were infected with the virus. Levels of the different immunoglobulin classes were correlated with viral RNA circulating in plasma or oral fluids and the proviral burden as detected by RT-PCR. Significant differences were determined between mothers and carrier children for immunoglobulin levels (p=0.037) and proviral burden (p=0.002). The overall estimate of IgG in plasma and sIgA in oral fluids could be correlated with the circulation of free viral RNA in both fluids and high proviral burden, and associated with HAM/TSP mothers. The detection of anti- tax IgG in plasma revealed differences between HAM/TSP mothers and their offspring. The study of immunological and molecular variables permitted the analysis of HTLV-1 circulation among families of Tumaco. The strong correlation between levels of IgM specific for the virus and viral RNA circulating in fluids indirectly confirmed the transmission of HTLV-1 among families.

The use of ELISAs for monitoring exposure of pig herds to Brachyspira hyodysenteriae

BackgroundSwine dysentery (SD), a mucohaemorrhagic diarrhoeal disease of pigs, results from infection of the large intestine with the spirochaete Brachyspira hyodysenteriae. ELISA systems using whole spirochaete cells (WC) and the B. hyodysenteriae outer membrane lipoprotein Bhlp29.7 previously have been established as potential diagnostic tools for SD. However, their true value in identifying infected herds remains unclear. The present study aimed to compare the performance of whole-cell and Bhlp29.7 based ELISAs in detecting specific immunoglobulin class IgG and IgM to B. hyodysenteriae in growing pigs, and additionally evaluated whether meat juice could serve as a source of specific antibodies.ResultsLevels of circulating IgG and IgM reacting with WC spirochaete preparations and recombinant Bhlp29.7 peaked 4-6 weeks post-infection in the experimentally challenged pigs, and remained elevated in the present study. In a cohort of pigs on an infected farm levels of antibody directed against both antigens showed a progressive increase with time. However, other than for the level of IgG against WC antigen, a significant increase in antibody levels also was observed in a cohort of pigs on a non-infected farm. In addition, assays using meat juice had 100% specificity and equivalent sensitivity to those based on serum, and likewise the best performance was achieved using the WC IgG ELISA.ConclusionsIgG ELISAs using either WC or Bhlp29.7 as plate-coating antigens were shown to be useful for monitoring the dynamics of B. hyodysenteriae infection in grower pigs. Of the two antigens, the WC preparation tended to give better discrimination between pigs from infected and non-infected farms. Testing of meat juice was shown to have potential for identifying infected herds.

Multiple protein extract microarray for profiling human food-specific immunoglobulins A, M, G and E

Existing food immunoglobulin (Ig) tests require large volumes of serum, are limited to one immunoglobulin class, are not amenable to high throughput analysis and only give a limited picture of the immunological response to food antigens. Conversely a new generation of Component Resolved Diagnostic systems using pure proteins is highly specific and totally dependent on the availability of the protein in its recombinant or natural origin form. Here we demonstrate a proof-of-concept of a microarray test based on protein extracts of food components. Our approach relies on innovations on three different fronts: the novelty of using arrayed food samples sequentially extracted with detergent and chaotropic agents, the ability to measure four different Ig classes simultaneously and the ability to analyse the generated data via a suitable bioinformatics/statistical analysis interface. This approach combines high numerical power of microarrays with automation, high throughput analysis and enables detailed investigation of the Ig profiles to food antigens. The prototype shown contains extracts of approximately 350 food ingredients that cover most of the food products found in the UK. Here we showed that the use of a sequential extraction technique to solubilise and then denature food samples has its benefits in the assessment of variations in antigenicity when tested with human sera. A patient dependent degree of class specificity was observed with human sera (IgG specificity correlates well with IgA > IgM >>>>> IgE). Besides generating a simultaneous profile for IgA, IgM, IgG and IgE the array system has shown good discrimination between challenge responders in atopic and non-atopic individuals. Poly- and mono-specific IgE responders were easily identified. The mathematical modelling of specific IgE content showed good correlations when compared with established IgE antibody testing assay (UniCAP). Although in its proof-of-principle stages, the immune profiling technique described here has the potential to provide unique insights into exposure/sensitization and establish relationships between specific immunoglobulin classes and subclasses against food protein antigens. In further developments, the immune profiling technique could also be extended to other related areas such as parasite and bacterial gut infection. Full analyses of large longitudinal and retrospective clinical trials are on going to determine the positive and negative predictive values of the technique.

Resultados do controle de qualidade de produtos hemoderivados: análise sanitária

The technical scientific unit of the National Institute of Quality Control in Health (INCQS), part of the Oswaldo Cruz Foundation, has a role of investigational quality control of products and services related to health surveillance, in particular medicines denominated plasma derivative products. This paper is based on the analysis of the 3100 plasma derivative products from January 2000 to December 2004: 31.6% (n=980) of human albumin, 28.7% (n=890) of factor VIII, 21.4% (n=662) of human immunoglobulins, 8.3% (n=257) of factor IX, 7.1% (n=22) of specific immunoglobulin classes containing anti-Rho (D) immunoglobulin, anti-hepatitis B, anti-tetanus, anti-rabies and anti-varicella-zoster and 2.91% (n=91) of prothrombin complex. The products submitted to analysis came from airports and frontiers of Brasilia, Rio de Janeiro and Sao Paulo, and products confiscated by the states of Pernambuco, Santa Catarina, Rio de Janeiro and Rio Grande do Sul. The type of analysis was characterized as: 92.3% control analysis; 5.9% fiscal analysis; 1.4% guidance and 0.4% preliminary analysis. In respect to imported plasma derivatives, 40.0% originated from the airport in Brasilia, 26.9% in Sao Paulo and 25.2% in Rio de Janeiro. In conclusion, 99.1% of the plasma derivative products analyzed was considered satisfactory and 0.9% unsatisfactory as identified by visual inspection, solubility, stability and chemical assays and pyrogenic and unspecific toxicity tests. Thus, the assessment of the quality of plasma derivative products is an essential tool for health surveillance.

Diagnosis of lyme borreliosis.

A large amount of knowledge has been acquired since the original descriptions of Lyme borreliosis (LB) and of its causative agent, Borrelia burgdorferi sensu stricto. The complexity of the organism and the variations in the clinical manifestations of LB caused by the different B. burgdorferi sensu lato species were not then anticipated. Considerable improvement has been achieved in detection of B. burgdorferi sensu lato by culture, particularly of blood specimens during early stages of disease. Culturing plasma and increasing the volume of material cultured have accomplished this. Further improvements might be obtained if molecular methods are used for detection of growth in culture and if culture methods are automated. Unfortunately, culture is insensitive in extracutaneous manifestations of LB. PCR and culture have high sensitivity on skin samples of patients with EM whose diagnosis is based mostly on clinical recognition of the lesion. PCR on material obtained from extracutaneous sites is in general of low sensitivity, with the exception of synovial fluid. PCR on synovial fluid has shown a sensitivity of up to >90% (when using four different primer sets) in patients with untreated or partially treated Lyme arthritis, making it a helpful confirmatory test in these patients. Currently, the best use of PCR is for confirmation of the clinical diagnosis of suspected Lyme arthritis in patients who are IgG immunoblot positive. PCR should not be used as the sole laboratory modality to support a clinical diagnosis of extracutaneous LB. PCR positivity in seronegative patients suspected of having late manifestations of LB most likely represents a false-positive result. Because of difficulties in direct methods of detection, laboratory tests currently in use are mainly those detecting antibodies to B. burgdorferi sensu lato. Tests used to detect antibodies to B. burgdorferi sensu lato have evolved from the initial formats as more knowledge on the immunodominant antigens has been collected. The recommendation for two-tier testing was an attempt to standardize testing and improve specificity in the United States. First-tier assays using whole-cell sonicates of B. burgdorferi sensu lato need to be standardized in terms of antigen composition and detection threshold of specific immunoglobulin classes. The search for improved serologic tests has stimulated the development of recombinant protein antigens and the synthesis of specific peptides from immunodominant antigens. The use of these materials alone or in combination as the source of antigen in a single-tier immunoassay may someday replace the currently recommended two-tier testing strategy. Evaluation of these assays is currently being done, and there is evidence that certain of these antigens may be broadly cross-reactive with the B. burgdorferi sensu lato species causing LB in Europe.

Analysis of SEVA TB ES-31 antigen specific immunoglobulins IgM, IgA and IgG in sera of sputum and culture positive pulmonary tuberculosis

Tuberculosis remains major health problem in India and developing countries Immunodiagnosis has important role in screening, diagnosis and management of tuberculosis. SEVA TB ES-31 antigen has shown potential in detecting tuberculous IgG antibody in earlier studies from our laboratory. In the present study we have analysedSEVA TB ES-31 antigen specific immunoglobulinsIgM, IgA and IgG in clinically and bacteriologically confirmed pulmonary tuberculosis cases to determine the usefulness of specific immunoglobulin class in the diagnosis of patients attending the hospital.Of the 30 cases of pulmonary tuberculosis 25 (83.3%) were positive for IgG, 19 (63.3%) for IgM and 16 (53.3%) for IgA. On combining IgG and IgM positivity, sensitivity was increased to 93.3%. While combining IgG and IgA positivity, sensitivity increased to 90%. However specificity was decreased to 66.6% and 70% for both of these combinations respectively. It could be envisaged from this study that IgG antibody detection against ES-31 antigen showed acceptable sensitivity (83.3%) and specificity (86.6%) compared to IgM or IgA alone or in combination. When immune responses were analysed according to degree of sputum positivity, IgG response was observed to be predominant in all grades, compared to IgM or IgA antibody. The addition of IgM or IgA as an adjunct test increases the sensitivity but at the cost of specificity. Hence the detection of IgG alone is more useful compared to IgM or IgA assay, in detecting tuberculosis disease cases coming to the hospital.

Evaluation of Antibody Class in Response to Bovine Collagen Treatment in Patients With Urinary Incontinence

Purpose Contigen(R) Bard(R) Collagen Implant (CI), made of highly purified bovine dermal type I collagen (BDC), is used as a bulking agent for the treatment of urinary stress incontinence. The humoral immune response to placement of this material in the urinary sphincter was evaluated. Materials and Methods In a prospective clinical study, patients were treated with CI in the urinary sphincter, and blood was collected at various timepoints following injection. Approximately 28 percent of the patients treated with BDC demonstrated specific antibodies against bovine type I collagen. Serum samples from 27 patients from this cohort were evaluated. The class specificity of circulating antibodies against bovine collagen was characterized by an indirect enzyme-linked immunosorbent assay. Results In all patients demonstrating an antibody response to bovine collagen, the predominant immunoglobulin class was IgG, found in 100 percent of sera samples. Immunoglobulin A was produced in approximately 40 percent of these patients, and IgM was seen in approximately 0.6 percent. No specific IgE was detected against bovine collagen in any serum sample. The highest concentrations of IgG and IgA antibody classes were observed 4 to 5 months after the initial treatment with CI. In the multicenter clinical trial, adverse events were reported in approximately 40 percent of all patients treated with CI (19). There was no correlation found between the production of a specific immunoglobulin class and the onset of any clinical adverse events. Conclusions In all sera from patients treated with CI for urinary stress incontinence, antibodies to bovine dermal collagen always were predominantly IgG. Immunoglobulin A was seen in less than half of the sera samples, and IgE was not observed. In addition, no change in the humoral response to CI over time was noted in patients demonstrating presensitization to bovine dermal collagen at the time of initial treatment. Clinical adverse events reported for patients demonstrating pretreatment antibodies against bovine dermal collagen did not differ in type or number when compared with patients having no presensitization.

Multiple successive immunoprinting: A fast blotting technique of a single agarose isoelectric focusing gel

Multiple successive pressure blottings of a single agarose isoelectric focusing gel were performed on normal and CNBr-activated nitrocellulose (NC) filters. The results obtained by multiple successive 10s immunoprints were compared to those obtained by a single 10 min immunoprint. To quantify the transfer efficiency of these techniques, a defined amount of radioactive material was separated by isoelectric focusing on agarose gel. After separation and pressure blottings of the gel the NC filters were submitted to autoradiography. The amount of radioactive material bound to the NC filters was determined by scintillation technique. The single 10 min pressure blot was more efficient than each of the multiple successive 10s prints. However, the latter procedure allowed equal resolution and resulted in a higher recovery of total radioactivity than the single immunoprint technique. The aim of this paper is to show how to obtain highly reproducible prints of electrophoretic patterns in an agarose gel of heterogeneous samples when accurate multiple immunodetections are to be performed. This technique was tested to characterize the grass pollen specific immunoglobulin classes and subclasses from an allergic patient.

Differential recognition of two cloned Brugia malayi antigens by antibody class

The humoral and cellular immune response to filarial parasites is complex. Numerous studies have shown that antibodies to a large number of protein and non-protein antigens may be produced over the course of infection and that immune recognition of any given antigen may vary by disease manifestation and by immunoglobulin class. We have used the techniques of molecular cloning to attempt to dissect this complex interaction, and describe here two clones, isolated from an expression library constructed from Brugia malayi genomic DNA, whose products are recognized by distinct immunoglobulin classes. A λgt11 fusion protein containing part of the B. malayi myosin tail region is recognized by antibodies of the IgG class from a high percentage of bancroftian filariasis patients. A fusion protein containing a collagen-like sequence is less frequently and weakly recognized under the same experimental conditions, but is almost universally recognized when the developing reagent is specific for IgE. We thus identify specific filarial proteins against which the infected human host responds preferentially with antibodies of a specific immunoglobulin class.

Importance of competitive binding in the detection of antigen specific bovine isotypes and subisotypes by the micro ELISA

An enzyme-linked immunosorbent assay was developed to detect and quantify the specific bovine immunoglobulin class response to Trypanosoma theileri, Dictyocaulus viviparus and infectious bovine rhinotracheitis virus. Comparative measurement of the specific immunoglobulin classes in whole serum was achieved using monospecific rabbit antibovine IgG1, IgG2 and IgM, followed by a goat anti-rabbit Ig-enzyme conjugate (antiglobulin ELISA). The results obtained in the antiglobulin ELISA compared favourably with the standard (or indirect) ELISA using the purified immunoglobulins. Competitive inhibition between specific immunoglobulins of different isotypes and subisotypes was the major disadvantage of the antiglobulin ELISA. This latter assay failed to detect specific IgG2 against T theileri antigen in both calf and adult whole serum. The inability to detect the specific IgG2 was a result of competitive inhibition by specific IgG1. However, competitive inhibition between specific immunoglobulins was not observed in either of the other test systems using D viviparus and infectious bovine rhinotracheitis virus antigens.

A simple method for detecting cells producing antibodies of specific immunoglobulin classes in the chicken

A simple plaque method for detecting cells producing antibodies of specific immunoglobulin class in the chicken is described. This method is based upon the inability of chicken antibody to activate guinea-pig complement. IgM- and IgG-specific plaque-forming cells in the spleen of chickens immunized with sheep red blood cells were detected using guinea-pig complement and rabbit anti-chicken-μ- or γ-chain serum. The specificity of the immunoglobulin class of plaques was confirmed by the abolition of class-specific hemolytic plaques after treatment of the lymphoid cells with rabbit antisera specific for chicken heavy chains in the presence of guinea-pig complement. The method is very simple and rapid, and IgM and IgG specific plaques are reliably detected.

Demonstration of extracellular immunoproteins in formalin-fixed renal biopsy specimens

Immunohistochemical techniques have been used for many years to establish the pathogenesis of renal disease. Deposits of immunoglobulin and complement are present in granules in glomerular capillary walls and mesangium in immune-complex disorders, whereas linear deposits of immunoglobulin and complement are found within the capillary walls in disorders due to antibodies directed against the glomerular basement membrane [1]. The diagnosis and classification of renal disease is often based on the detection of specific immunoglobulin classes and their site of deposition [2]. Either immunofluorescent or immunoperoxidase methods can be applied to renal tissue that has been frozen at the time of biopsy, and the two methods give similar results [3–5]. In almost every case, these methods are unable to demonstrate extracellular deposits of immunoglobulin and complement if the tissue has been fixed in formalin. Following the reports of Huang, Minnasian, and More [6] and of Curran and Gregory [7], we have investigated the use of an immunoperoxidase technique on formalin-fixed wax-embedded needle-biopsy specimens of the kidney after prior treatment with trypsin. We have used the unlabeled antibody, peroxidase-antiper-oxidase (PAP) method of Sternberger [8] and have shown that controlled treatment with trypsin prior to the PAP method enables extracellular deposits of both immunoglobulin and complement to be demonstrated. The results are compared with those obtained from standard immunofluorescent methods performed on renal tissue frozen at the time of biopsy.

Penicillin antibodies and immunoglobulin production in newborn infants: studies with chemically modified bacteriophage.

Extract: With the use of the highly sensitive technique of inactivation by antibody of growth of bacteriophage to which a hapten penicillin is chemically coupled, we found anti-penicillin antibodies (APA) in the sera of all 25 mothers examined and their 27 newborn infants. Precise quantitation of antibodies was not performed in this study, but with the use of optimum dilution of the sera for antibody detection as the indicator, the efficiency, (i.e., a function depending both on amount and on affinity) of antibodies in the sera of the mothers was higher by a factor of 5–10 when compared with the sera of the neonates. The immunoglobulin classes of APA were determined by both the direct method after removal of a single immunoglobulin class by immuno-adsorption on a Sepharose-antiimmunoglobulin column, and by indirect methods such as destruction of heat-labile immunoglobulin (immunoglobulin E is heat labile), and breakdown of macroglobulin (IgM) with 2-mercaptoethanol (2-ME). There was good correlation between the results by both methods, although occasionally some discrepancies were found. Most sera had APA of human immunoglobulins G, M, A, and E (IgG, IgM, IgA, and IgE) classes, but none of the sera of seven mothers and five infants contained IgD APA. More infants than mothers had a high proportion of IgM APA, whereas there were no significant differences between the two groups with regard to IgG, IgA, and IgE APA. In premature infants, all four immunoglobulin classes of APA were found, the youngest infant examined being 6 months gestational age. The proportion of each APA immunoglobulin class in paired mother-infant sera was approximately similar in most cases. However, several pairs were found to have marked discrepancies in the proportion of a specific immunoglobulin class. In three pairs, IgA antibodies were present in mothers' sera diluted 1/500–1/1,000 but not in the infants' sera diluted 1/50–1/100 when other immunoglobulin classes of APA were present. In several pairs, the mother had no IgE APA, whereas it was present in the infant, and, conversely, some mothers had IgE APA, whereas their infants had none. No such remarkable discrepancies were found in the IgG and IgM classes of APA. In the two sets of twins, marked differences were found in each infant pair in the proportion of immunoglobulin class APA. These results indicate that the fetus is able to synthesize IgE antibodies, and are highly suggestive of independent synthesis of IgA, IgM, and IgG antibodies in mother and fetus. These results also suggest that synthesis of the various immunoglobulins by the fetus occurs as early as 6 months gestational age, if not earlier, and that IgE and IgA probably do not pass through the placenta consistently. The presence of antipenicillin antibodies in infants whose mothers had no history of receiving penicillin or its derivatives during pregnancy leads one to speculate that penicillin may be a pollutant of the food we eat and drink. However, the possibility that antibodies cross-reacting with penicillin are present in fetuses must be taken into consideration, particularly as our studies have shown that all neonates have antibodies of various immunoglobulin classes with specificity directed towards the dinitrophenyl hapten.Speculation: Evidence that the immune system of the human fetus is competent and responds to antigenic stimulation in utero indicates that possibilities exist to immunize infants prenatally in order to supply them with adequate and specific active antibodies at birth. The presence of antibodies to penicillin in the sera of all newborn infants studied indicates either that penicillin is a universally present antigen, possibly as a pollutant passing from mother to fetus, or that cross-reacting antibodies to penicillin exist in the fetus.

Phylogeny of hypersensitivity. I. anaphylactic responsiveness of the frog, Rana pipiens

Rana pipiens immunized with Salmonella typhosa vaccine developed (anaphylactic) shock reactions on challenge with corresponding soluble bacterial antigen (SBA) but not with washed organisms. Hemagglutinating serum antibody to lipopolysaccharide (LPS) and anti-H agglutinins were demonstrated in responsive frogs. Shock reactions were not influenced by hibernation at 4° C. or by total aqueous immersion. Less uniform results were produced by challenging injections of bovine gamma globulin in 6 of 36 frogs that developed corresponding 2-mercaptoethanol (2-ME)-sensitive hemagglutinating antibodies and 2-ME-resistant precipitins following effective immunization with bovine gamma globulin (BGG) added to typhoid vaccine. Rabbit antiserum to S. typhosa containing anti-O and anti-H agglutinins, 2-ME-sensitive hemagglutinating antibody to LPS, and 2-ME-resistant precipitins to SBA passively transferred anaphylactic responsiveness to the frog, but this could not be effected with rabbit antiserum to BGG. Identification of a shock organ or pathologic changes could not be demonstrated in affected animals. The frog, an ectothermic amphibian representing a transitional position on the phylogenetic scale in production of specific immunoglobulin classes, may develop hypersensitivity manifestations under delineated conditions of exposure and challenge to a microbial antigen .

Identification of IgA in Rat Serum and Secretions

Abstract Although serum and/or secretory IgA has been well defined in the human as well as in several laboratory animals including mice, rabbits and dogs (1, 2), this immunoglobulin has not been adequately identified and characterized in the rat. Several reports have appeared in which a rat serum immunoglobulin of relatively slow mobility was referred to as IgA (3–5) or γ1 (6). This identification was made, for the most part, on the basis of electrophoretic mobility. The purpose of this communication is to demonstrate that the mobility of rat serum IgA is faster than that of the presently known rat immunoglobulins, and that some of the previous papers dealing with rat IgA refer to a serum immunoglobulin which may in fact be homologous to mouse IgG1 (IgF). Of the many possible criteria which can be used in identifying a specific immunoglobulin class within a species, the most precise would appear to be a comparison of amino acid sequences between known immunoglobulins and the protein being studied.

Assignment of Direct and Facilitated Hemolytic Plaques in Mice to Specific Immunoglobulin Classes

Summary The kinetics of appearance of hemolytic plaque-forming cells in immunized mouse spleens has been studied with the aid of immunoglobulin specific antisera and myeloma protein inhibitors. “Facilitated” or “indirect” plaques were developed with antisera specific for γ1, γ2 and γM, and for κ light chains. Facilitation with anti-γM antiserum suggests the presence of two classes of mouse macroglobulin, one responsible for direct plaques and the other requiring an anti-globulin reagent in order to produce lysis. The previously held concept that all indirect plaques represent “7 S” antibody is no longer tenable.