Porphyromonas gingivalis-specific serum IgG and IgA antibodies originate from immunoglobulin-secreting cells in inflamed gingiva.

Patients with adult periodontitis (AP) exhibit elevated serum antibody levels to Porphyromonas (Bacteroides) gingivalis; however, it is not known whether these antibodies originate from plasma cells in the local disease site or from peripheral lymphoid tissues. We studied the isotype and subclass levels and origin of antibodies to P. gingivalis fimbriae, since elevated serum anti-fimbriae responses were seen when compared with sera of healthy controls. IgG anti-fibriae titres were dominant and the subclass response was IgG3 much greater than IgG1 greater than IgG2 much greater than IgG4; however, some IgA anti-fimbriae antibodies were also seen. The IgA subclass fimbriae-specific response was mainly IgA1; however, significant IgA2 anti-fimbrae antibodies were seen. We also assessed numbers of anti-fimbriae antibody producing cells from peripheral blood mononuclear cells (PMBC) and from either healthy or inflamed gingiva of AP subjects. Gingival mononuclear cells (GMC) of AP patients exhibited high numbers of immunoglobulin-producing (spot-forming) cells (SFC) including fimbriae-specific antibody secreting cells in a pattern of IgG greater than IgA greater than greater than greater than IgM. However, low numbers of SFC were seen in GMC from healthy gingiva; further, no anti-fimbriae SFC responses were noted in healthy GMC. Although no fimbriae-specific immunoglobulin-producing cells were seen in PBMC, low numbers of antigen-specific SFC were found in pokeweed mitogen-triggered PBMC from AP subjects. Treatment of AP patients for plaque and surgical removal of inflamed gingiva resulted in significant reductions in serum anti-fimbriae responses. These studies show that AP patients exhibit brisk serum IgG and IgA subclass anti-fimbriae antibodies, whose origin appear to be the plasma cells present in the localized inflamed tissues.     

The predominance of IgG1 and IgG3 subclass antisperm antibodies in infertile patients with serum antisperm antibodies.

Mouse monoclonal antibodies (MAb) specific for each of the four human IgG subclasses and immunofluorescence flow cytometry were used to evaluate the subclass of the IgG antibody response to sperm in serum samples from 13 men and 6 women with a high titer (greater than 1:15,625) of IgG antisperm antibodies (ASA] determined by an indirect immunobead test. Five sera without ASA were also studied as a control. All 19 (100%) of the ASA-positive sera contained immunoglobulin (Ig)G ASA of the IgG1 and IgG3 subclasses. A 1:1 correlation was observed between the presence of IgG1 and IgG3 ASA. IgG2 was essentially undetectable, while IgG4 reactivity, although less intense than IgG1 and IgG3, was more prominent in the sera from the five vasectomized men. The ability of the IgG1 and IgG3 ASA-positive sera to deposit complement (C) on sperm was demonstrated by the concomitant binding to antibody-laden sperm of polyclonal antibodies to the membrane attack complex (C5b-9) of C. Both C-fixing and non-C-fixing ASA-positive sera were found to possess IgG1 and IgG3 antisperm antibodies. The predominance of IgG1 and IgG3 subclasses suggested a T-cell dependent immune response to sperm antigens.     

IgG subclass distribution of thyroglobulin antibodies in patients with thyroid disease

The IgG subclass distribution of thyroglobulin antibodies (TgAb) has been studied in Hashimoto and Graves’ patients by several investigators with conflicting results, in part explainable by methodological problems. We have recently developed a quantitative ELISA to measure in absolute terms the serum concentration of TgAb subclasses. The aim of the present study was to apply this method in a large series of patients with autoimmune as well as, for the first time, non-autoimmune thyroid diseases. We examined 28 patients with Hashimoto's thyroiditis, 30 with Graves’ disease, 21 with thyroid carcinoma and 18 with non-toxic goitre, all selected for the presence of TgAbs. The results indicated that TgAbs in thyroid diseases were not restricted to any particular isotype, but comprised all four IgG subclasses. IgG1 was represented similarly in the four groups. The same was true for IgG3, even though its contribution to the total antibody content was very small. IgG4 was the dominant subclass in patients with Graves’ disease, thyroid carcinoma and non-toxic goitre, probably reflecting a prolonged antigenic challenge. In Hashimoto's thyroiditis IgG2 was dominant, possibly because T helper lymphocytes infiltrating the thyroid are typically Th1 type.     

Severity of infections in IgA deficiency: correlation with decreased serum antibodies to pneumococcal polysaccharides and decreased serum IgG2 and/or IgG4

In order to define abnormalities of humoral immunity which determine susceptibility to respiratory tract infections in IgA-deficient adults, serum IgG subclass concentrations, and serum concentrations of pneumococcal antibodies and Haemophilus influenzae type B (Hib) antibodies sera from IgA-deficient adults with and without susceptibility to respiratory tract infections were compared. Infection susceptibility was not related to the degree of IgA deficiency, but was related to deficiency of IgG4 and, to a lesser extent, IgG2, as well as to low basal serum concentrations of pneumococcal polysaccharide antibodies. The combination of IgG2 and/or IgG4 deficiency and a non-protective basal serum concentration of antibody against two or more pneumococcal polysaccharides was present in the serum of six of 12 (50%) patients with severe infections, but only one of 44 (2%) patients without infections. Furthermore, the preservation of antibody responses against the most immunogenic pneumococcal polysaccharide type 3, but not against the less immunogenic types 7F, 9N and 14, in patients with severe infections suggested that abnormalities of pneumococcal polysaccharide antibody responses might include defects of affinity maturation.     

Specificity of IgG subclass antibodies in different clinical manifestations of leprosy.

We analysed specific IgG subclasses levels to Mycobacterium leprae sonicate extract (MSE), lipoarabinomannan B (LAM) and phenolic glycolipid I (PGL-I) in the sera of leprosy patients with different clinical manifestations. IgG2 was found to be the predominant antibody to MSE regardless of clinical manifestations, and IgG1 response was mostly seen in lepromatous patients. IgG3 reacted only rarely but IgG4 reacted relatively more in certain clinical groups such as borderline lepromatous and lepromatous with erythema nodosum leprosum (ENL) reaction. Most of the IgG subclass responses to MSE could be accounted for reactivity with LAM, suggesting that LAM is the major immunogen involved in the pathogenesis of leprosy. In contrast to LAM, PGL-I antigen showed considerably lower reactivities for IgG subclasses. An association between IgG subclass responses and clinical manifestations of leprosy was also seen. Whereas borderline lepromatous patients were found to have significantly higher levels of IgG2 and IgG4 to MSE, lepromatous patients had elevated levels of IgG1 and lower levels of IgG2. An interesting observation, however, was the significantly higher levels of IgG2 to LAM in the pure neuritic leprosy patients.     

IgG subclass antibodies against Parietaria judaica in normal and allergic subjects

IgG antibody response to the inhalant allergen Parietaria judaica (Pj) and IgG subclass distribution were studied in 82 normal subjects, divided into three groups according to age (0–1, 1–20, and 20–60 years) and in 32 allergic subjects aged 20–60 years. Both normal and allergic subjects showed an IgG response, and all had IgG1 antibodies specific for PjE. Serum IgG2, IgG3, and IgG4 against PjE were detectable in 36%, 46%, and 22% of normal subjects, and in 58%, 31%, and 65% of allergic subjects, respectively. A significant difference in class distribution between allergic and age-matched normal subjects was found only for IgG4 antibodies against PjE (65% and 17%; P<0.01). The ELISA results were also analyzed quantitatively, taking into account the relative proportion of specific antibodies. Thus, in normal subjects IgG1 antibodies showed a decreasing trend as the age rose, while no differences according to the age of the subject were found for IgG2 and IgG4. When data from allergic subjects (20–60 years) and the age-matched normal group were compared, they were different for the relative percentage of IgG2 only, showing for this a significantly lower value (P< 0.001). The present data indicate that normal and allergic subjects show differences in the IgG isotype distribution depending on their sensitivity and duration of allergen exposure.     

Clinical significance of IgG subclass antibodies to wheat flour antigens in bakers

We measured the IgG subclass antibody levels to wheat flour in 42 bakers and 20 controls with an enzyme immunoassay. The levels of total IgG, IgG1 IgG2 and IgG4 antibodies were significantly higher in the bakers than in the unexposed controls. The presence of anti-wheat flour IgG subclass antibodies in the bakers was correlated with various clinical variables including IgE levels, duration of asthmatic or rhinitis symptoms, skin prick test response, peripheral blood eosinophil levels, bronchial histamine reactivity and responses to nasal challenge with wheat flour. The IgG subclass antibody levels of the total cohort of bakers did not correlate with any of the measured clinical variables. However, among men specific IgG4 and IgG1 antibody levels correlated negatively with total IgE levels and duration of rhinitis, respectively. We conclude that IgG and IgG subclass levels to wheat flour in bakers reflect exposure, but that it is not related to any specific clinical situation. The exact pathogenic role of these antibodies in the development of occupational asthma and rhinitis is thus not clear.     

Cytomegalovirus IgG and IgA serum antibodies in a study of HIV infection and HIV related diseases in homosexual men.

The significance of serum IgG and IgA antibodies to cytomegalovirus (CMV) at various stages of human immune deficiency virus (HIV) infection was studied in 175 homosexual men. Sera were obtained from 123 HIV seropositives [41 asymptomatic, 29 with lymphadenopathy associated syndrome (LAS), 22 with AIDS related complex (ARC), and 31 AIDS patients], 17 HIV seroconverters, and 35 HIV asymptomatic seronegatives. The sera were tested blindly for CMV IgA and IgG antibodies using the immunoperoxidase assay (IPA) and CMV infected human embryo cells. Cross-sectional analysis of CMV IgG antibodies at a titer of greater than or equal to 20 showed 87% and 100% prevalence in the HIV seronegative groups and in the HIV seropositive groups, respectively (P less than 0.05). CMV IgG antibodies at a titer of greater than or equal to 80 were present in significantly higher proportions among the HIV seropositive subjects of the various groups as compared with the HIV seronegative homosexual men. However, in the HIV seronegatives who later seroconverted to HIV, a significantly higher prevalence of CMV antibodies (35%) was detected before HIV seroconversion, as compared with the persistently HIV seronegative subjects (14.3%) (P less than 0.05). The HIV seronegatives pre-HIV seroconversion also exhibited a significantly higher geometric mean titer (GMT) of CMV IgG antibodies (62.17 +/- 0.64) as compared with the persistently HIV seronegatives (34.0 +/- 0.6) (P = 0.03). Significantly higher GMTs of CMV IgG antibodies were detected in all the HIV seropositive groups as compared with the persistently HIV seronegative group. CMV IgG antibodies were not detected in the HIV seronegative subjects.(ABSTRACT TRUNCATED AT 250 WORDS)     

Bindine affinities of allergens from pollen, mites, and house dust for specific IgG subclass antibodies

The distribution and affinity of IgG subclasses against various aeroallergens were assessed by inhibition of specific antibody binding. Two parameters from the dose-response curves were taken as indicative of antibody affinity: the point of 50% inhibition and the value of the slopes on double-log plots. It was found that IgG4 antibody specific for aeroallergens (i.e., from pollens of several species of Gramineae, Olea europaea , and Parietaria judaica and from house dust) usually exhibits high affinity, except for Dermatophagoides pteronyssinus . High binding affinity was also displayed by IgGl subclass antibodies against the allergens of O. europaea and P. judaica . Distinct IgG subclass affinity profiles were observed for the allergens of grass pollen (i.e., Holcus lanatus ) and dust mites (i.e., D. pteronyssinus ). These results demonstrate that IgG subclass distribution, as well as antibody affinity, depends on the nature of the sensitizing allergen.     

Enzyme-linked immunosorbent assay for subclass distribution of human IgG and IgA antigen-specific antibodies

In this study we describe an ELISA using monoclonal antibodies to IgG 1, 2, 3, 4, IgA1 and IgA2 for determining the subclass distribution of human-specific antibodies. No cross-reactivity of the subclass-specific reagents under the conditions used was observed. The sensitivity was 0.5 ng/ml for IgG1, 3, 4; 1.5 ng/ml for IgG2 and 50 ng/ml for IgA1 and IgA2. The reproducibility as described by the coefficient of variation calculated on repeated runs was 8-26% if the data were obtained by relating the absorbance values to a positive serum run in the assay, 17-58% when relating the OD figures to those of a standard myeloma plate. The method may be considered semiquantitative with high sensitivity and specificity, easy to handle and with small day-to-day variation. The assay has been applied to a number of antigens of protein and polysaccharide nature.     

Prospective estimation of IgG, IgG subclass and IgE antibodies to dietary proteins in infants with cow milk allergy

Prospectively, serum levels of IgE, specific IgE antibodies (AB) to whole cow milk protein (CMP), bovine se-albumin, bovine immunoglobulin, bovine lactoferrin, bovine lactalbumin and beta-lactoglobulin (BLG), IgG and IgG subclass antibodies to ovalbumin (OA) and BLG, and IgG4 RAST to CMP (bovine whey) were measured in 39 infants with cow milk protein allergy (CMPA) at birth (cord blood), at time of diagnosis and before and after milk challenge at the age of 12 months. Immunological measurements were also undertaken in 33 control infants without CMPA at birth, at 6 months and at 18 months. At no time, were differences found between the levels of IgG and IgG subclass AB to OA and BLG in control versus infants with CMPA. In the 39 infants with CMPA no correlation was found between the levels of IgE, IgG and IgG subclass AB in cord blood and subsequent levels of these values, irrespective of the type of CMPA (IgE-mediated (CMA) or non-IgE-mediated (CMI)), and irrespective of whether remission had occurred. In cord blood 25/33 (76%) of the infants with CMPA had specific IgE-AB to one or more of the bovine milk proteins indicating a prenatal intrauterine sensitization to cow milk protein. At 6 months the frequency of specific IgE-AB to bovine milk proteins was significantly (p less than 0.05) higher in infants with CMA versus CMI, and at 12 months total serum-IgE and the increase of these specific IGE-AB and RAST to CMP were significantly higher (p less than 0.05) in infants with persistent CMA. From 6 to 12 months withholding milk resulted in a significant fall in specific IgE-AB to CMP, and IgG, IgG1 and IgG4 anti-BLG followed by an increase after milk challenge. Decreasing levels of IgG anti-OA from birth to 6 months reflect passive maternal transfer of IgG through the placenta, and increasing levels of IgG anti-BLG, already from birth to 6 months, may represent an early exposure to CMP in all infants. Significantly higher levels (p less than 0.05) of IgG anti-OA AB, IgG1 and IgG4 anti-BLG AB were found in infants with persistent CMA, indicating a close relation between the synthesis of IgE and IgG and between IgE and IgG subclasses (IgG1 and IgG4) in symptomatic cow milk-allergic individuals.(ABSTRACT TRUNCATED AT 400 WORDS)     

The significance of IgG subclasses and mannan-binding lectin (MBL) for susceptibility to infection in apparently healthy adults with IgA deficiency.

The aim of this study was to investigate the significance of IgG subclasses and MBL for susceptibility to infection in association with IgA deficiency. The study population consisted of 139 apparently healthy adult blood donors with IgA deficiency and normal serum levels of IgG and IgM, and an increased susceptibility to infection demonstrated at a population level. Additionally, 216 controls matched for age and sex were investigated. IgG4 deficiency was more common and the mean level of IgG4 lower in persons with IgA deficiency than in the controls. No significant associations could be demonstrated between overt IgG subclass deficiencies and increased susceptibility to infection. However, when the mean concentrations of IgG subclasses were analysed with regard to medical history, that of IgG1 was lower in persons who reported recurrent viral respiratory infections, that of IgG3 in persons who had episodes of severe infection in their history, and that of IgG4 in persons who had recurrent mild respiratory infections, compared with those who had no particular history of infections. In contrast, MBL deficiency-alone or combined with that of the IgG subclass-was not associated with increased susceptibility to infection in persons with IgA deficiency. The results indicate that the proneness to infections observed in a population of otherwise healthy persons with IgA deficiency can only for a small part be accounted for by concomitant deficiencies of IgG subclasses. Contrary to expectations, no synergism between the deficiencies of IgA and MBL could be demonstrated.     

A comparative study of IgG subclass antibodies in patients allergic to wasp or bee venom

IgG1 and IgG4 antivenom antibody responses were compared in groups of patients who had experienced systemic reactions to wasp ( Vespula spp.) or bee stings. Pretreatment serum IgG4 antibody levels were low in both groups, but IgG4 antibodies were significantly raised in bee-allergic patients ( P <0.002), probably reflecting their greater exposure to stings than wasp-reactive patients. No direct or indirect relationships were found, in untreated bee or wasp patients, between IgG1, IgG4, or IgE antibody levels and the severity of a patient's last systemic reaction to a sting. After a 12-week course of venom immunotherapy (VIT), IgG1 antibodies increased significantly only in wasp-sensitive patients ( P <0.001), although both groups responded with marked increases in venom-specific IgG4 ( P <0.01). Wasp-allergic subjects who responded to VIT with high production of specific IgG4 showed the greatest increases (pre- to post-VIT) in IgE antibodies (P<0.05). This group also demonstrated a direct correlation ( P <0.05) between post-VIT levels of IgE and IgG1 antibodies, a finding contrary to an IgE-immunoregulatory role for IgG 1. High levels of venom-specific IgG1 alone, or in combination with IgG4, were not protective in three patients who suffered repeated adverse reactions to bee VIT, showing that absolute levels of IgG subclass antivaenom antibodies are not reliably indicative of clinical responsiveness in individual patients.     

Differential binding of IgG and IgA antibodies to antigenic determinants of bovine serum albumin

The aim of this study was to investigate the recognition pattern of bovine serum albumin (BSA), a major dietary protein by serum IgG and IgA antibodies. Anti-BSA IgG and IgA antibodies were measured by ELISA technique in 3 different cohorts: 578 unselected persons, 84 new-onset insulin-dependent diabetes mellitus (IDDM) patients and 103 atopic persons. In order to characterize the recognition pattern of the different BSA domains, recombinant BSA and recombinant fragments covering the 3 BSA domains were produced. BSA digestion was monitored in simulated gastric fluid experiments by means of domain specific monoclonal antibodies. IgG and IgA antibody titres to native BSA were highest in IDDM patients. The three major BSA domains were equally well recognized by IgG antibodies of the three cohorts. Interestingly all three study groups showed a dissociation of their IgG and IgA antibody response to the first BSA domain. The ratio of IgG to IgA antibodies recognizing this domain was 93%/42% in controls, 92%/37% in IDDM patients and 80%/47% in atopic persons. In simulated gastric fluid experiments, the first BSA domain was the first to become undetectable to specific monoclonal antibodies during digestion. In conclusion humoral IgG and IgA antibodies recognize the major BSA domains with different frequencies. The N-terminal domain of BSA, the first to be degraded during simulated gastric digestion is less well recognized by IgA antibodies. This suggests that early digestion is negatively correlated to the IgA antibody response and that the IgA response associated to the gut associated lymphoid tissue (GALT) and the systemic IgG antibody responses are independent.     

Subclass distribution of IgA antibodies in saliva and serum after immunization with Haemophilus influenzae type b conjugate vaccines

IgA subclass distribution of antibodies against capsular polysaccharide (PS) of Haemophilus influenzae type b (Hib) was studied in saliva and serum samples of children vaccinated with two (n = 58) or three doses (n = 53) of Hib vaccine. One month after the second dose of Hib conjugate vaccine, at 7 months old, 40% of the children had IgA1 and 41% had IgA2 anti-Hib PS antibodies in saliva. One month after the third dose, at 15–25 months old, IgA1 was the predominating subclass; 72% of the children had IgA1, 26% had IgA2 anti-Hib PS in saliva. The mean concentration of IgA1 anti-Hib PS, expressed as optical density (OD) values, was significantly higher after three doses (OD 80.7) than after two doses (OD 18.9). The mean concentration of IgA2 did not change significantly after the third dose (OD 23.8 after two doses, OD 18.1 after three doses). In serum, IgA1 anti-Hib PS predominated both after two (17% had IgA1, none had IgA2) and three doses (72% had IgA1, 4% had IgA2) of Hib vaccine. In conclusion, both IgA1 and IgA2 anti-Hib PS were found in saliva of immunized children after two doses of Hib conjugate vaccine, whereas the third vaccine dose induced a shift towards IgA1 anti-Hib PS dominance in saliva.     

Subclasses of IgA antibodies in serum and saliva samples of newborns and infants immunized against rotavirus

Little is known about subclass levels of IgA in serum or saliva of infants in the perinatal period. We have previously shown that very young infants are capable of responding to an experimental rotavirus vaccine with both serum and salivary IgA, and that small amounts of IgA are already detectable in cord blood of these infants. In the present study, total IgA1 and IgA2 antibodies in serum and saliva samples of some of these infants at birth, at 6 weeks of age, and at 12 weeks of age, were determined by a quantitative ELISA. Also, subclass-specific IgA antibodies to the rotavirus group A common antigen were determined by ELISA. The ratio of average serum concentrations of IgA1 to IgA2 for 14 infants at 6 weeks of age was 19:1, while in saliva it was 5:1. Between 6 and 12 weeks of age levels of serum IgA1 increased by 25%, while levels of IgA2 did not increase perceptibly. Concentrations of IgA1 were higher in infant sera than in saliva, while concentrations of IgA2 were slightly higher in saliva than in serum. When calculated as specific ELISA units per mg IgA1, more salivary IgA1 was specific for rotavirus than serum IgA1. Further studies are needed to determine when infant IgA2 levels rise to values more characteristic of children and adults. This may be of significance for infant mucosal immunizations if secretory IgA2, more resistant to bacterial proteases than IgA1, is required for efficient defence of the respiratory and intestinal tracts.     

IgG subclass distribution of antibody responses to protein and polysaccharide mycobacterial antigens in leprosy and tuberculosis patients

Immunoenzymatic assays were developed for the measurement of antibodies against mycobacterial lipoarabinomannan (LAM), a cell-free proteic extract (CFX) of Mycobacterium leprae, and the 38-kD protein antigen of M. tuberculosis. Sera from 108 leprosy patients, belonging to all clinical–immunological forms of the spectrum, and 81 patients with localized or disseminated tuberculosis (TB) were tested for antibodies of the four IgG subclasses. Standard calibration curves were used to allow comparisons between results of different isotypes and specificities. Mean concentrations of total IgG antibodies were higher in the overall leprosy population than in TB patients. In leprosy, levels of anti-CFX increased from tuberculoid toward lepromatous forms, with a clear switch from IgG1 to IgG2 subclass predominance. A similar IgG1 to IgG2 conversion was observed in anti-LAM antibodies, although total levels of anti-LAM were similar in patients with tuberculoid and lepromatous forms. In TB, antibodies against polysaccharide and protein antigens were both predominantly of IgG1 subclass, whatever the patient's clinical status, although lower in disseminated forms, probably due to concomitant HIV infection. A hypergammaglobulinaemia was also found in most leprosy and TB patients. In TB this was due to increased IgG1 and IgG3, especially in HIV co-infected patients. Based on the current knowledge of the influence of T cell-secreted cytokines on human immunoglobulin isotype expression, these results do not fit with a putative role of Th1 (such as found in TB and tuberculoid leprosy (TT)) and Th2 (such as found in leprosy lepromatous (LL) leprosy) environment in the isotypy of antibody responses in mycobacterial infections. Nor do variations of isotypy according to pathological conditions seem to be related to the biochemical nature of antigens, since antibodies to LAM and protein antigens had comparable evolutions of their subclass distribution. Other factors are to be investigated in order to understand better the significance and possible roles of antibodies in mycobacterial diseases.     

The significance of blood levels of IgM,IgA, IgG and IgG subclasses in Sudanese visceral leishmaniasis patients

We developed an ELISA test using leishmania antigenic extracts to detect antigen-specific antibody responses, including subclass and isotype analysis, in visceral leishmaniasis (VL) patients from the Sudan. A total of 92 parasitologically proven patients were compared with cutaneous leishmaniasis, schistosomiasis, malaria, onchocerciasis and tuberculosis patients, as well as with healthy endemic and non-endemic controls. Some VL patients were examined before and after chemotherapy. VL patients showed significantly higher IgG responses compared with all other groups (93·4% sensitivity, 93·7% specificity), and higher (but not significantly) IgM responses. All groups showed low IgA levels. All groups showed low IgA levels. All IgG subclasses, IgG1, 2, 3, and 4, showed higher levels in patients than all other groups, with IgG1 and IgG3 levels being significantly reduced following treatment. The rank order for specificity and sensitivity for IgG subclasses was IgG3 > IgG I> IgG2> IgG4.     

IgG and IgA subclass mRNA-bearing plasma cells in periodontitis gingival tissue and immunoglobulin levels in the gingival crevicular fluid

The humoral immune response, especially the production of IgG and IgA, is considered to have a protective role in the pathogenesis of periodontal disease, but the precise mechanisms are still unknown. In order to determine local IgG and IgA production, we investigated the presence of human IgG and IgA subclass mRNA-bearing plasma cells within periodontal tissue by in situhybridization using digoxigenin-labelled oligonucleotide probes in 24 gingival biopsy samples (pocket depth>5 mm) which were obtained from eight patients with adult periodontitis. Furthermore, we examined IgG and IgA subclass proteins and digested IgA1 Fab portions in the gingival crevicular fluid (GCF), corresponding to the sites from which the tissues were taken, by ELISA. IgG and IgA subclass mRNA-expressing cells were detected in all serial formalin-fixed/paraffin-embedded gingival tissue sections sampled. Plasma cells showed strong cytoplasmic staining with a high contrast and a good retention of morphology with these probes. IgG1 mRNA-expressing cells were predominant (mean 63%) and IgG2 mRNA-expressing cells were present at around 23% of total IgG plasma cells, while IgG3 and IgG4 mRNA-expressing cells were present to a much lesser extent (3% and 10%, respectively). Similar proportions of IgG subclass proteins in GCF were detected, which were also consistent with ‘normal’ serum levels. In terms of IgA subclass, IgA1 mRNA-positive cells were predominant (mean 65.1%, P<0.001). In contrast, IgA2 protein in the GCF samples were detected at higher concentrations than IgA1 (P<0.001). The ratio of total IgG to IgA mRNA-positive plasma cells was ≈7.5:1. There was a good correlation between the amounts of IgG subclass proteins in GCF and the number of IgG subclass mRNA-positive cells in the same sites, but not between IgA subclass proteins and the number of IgA subclass mRNA-positive cells. These data suggest that IgG and IgA subclass proteins can be locally produced in the periodontitis gingiva. In addition, as we detected IgA1 Fab fragments in GCF, this is further confirmation that secreted IgA1 protein in GCF may be digested by periodontal bacteria.     

Solid-phase radioimmunoassay of IgA, IgG, and IgM antibodies to human rotavirus.

A solid-phase radioimmunoassay (RIA) has been developed for the detection of human rotavirus-specific IgA, IgG, and IgM antibodies. Nebraska calf diarrhea virus grown in LLC-MK2 cell cultures in the presence of trypsin was directly adsorbed onto polystyrene balls, and antibodies that attached to the virus-coated balls were detected by subsequent binding of 125I-labeled antibodies specific to human alpha, gamma or mu chains of human Iga, IgG, or IgM immunoglobulins. A total of 116 serum specimens from 58 adult patients were tested. Binding ratios between the positive and the negative serum varied between 5 and 15, occasionally being 20 or more in the IgA and IgG assays, but rarely exceeding 3 in the IgM assay. The RIA was found to be more sensitive in detecting antibodies to rotavirus than the complement fixation (CF) test, the RIA titers obtained being 50--100 times as high as the CF titers. The method described offers a possibility of evaluating the immune response to human rotavirus and of detecting recent infection.