Subclass Antibody Levels Research Articles

Immunoproteomics of cow's milk allergy in Mexican pediatric patients

Immunological mechanisms of non-IgE-mediated cow's milk protein allergy (CMPA) are not well understood. Such a circumstance requires attention with the aim of discovering new biomarkers that could lead to better diagnostic assays for early treatment. Here, we sought both to investigate the mechanism that underlies non-IgE-mediated CMPA and to identify cow's milk immunoreactive proteins in a Mexican pediatric patient group (n = 34). Hence, we determined the IgE and IgG1–4 subclass antibody levels against cow's milk proteins (CMP) by ELISA. Then, we performed 2D-Immunoblots using as first antibody immunoglobulins in the patients'serum that bound specifically against CMP together with CMP enrichment by ion-exchange chromatography. Immunoreactive proteins were identified by mass spectrometry-based proteomics. The serological test confirmed absence of specific IgE in the CMPA patients but showed significant increase in antigen-specific IgG1. Additionally, we identified 11 proteins that specifically bound to IgG1. We conclude that the detection of specific IgG1 together with an immunoproteomics approach is highly relevant to the understanding of CMPA's physiopathology and as a possible aid in making a prognosis since current evidence indicates IgG1 occurrence as an early signal of potential risk toward development of IgE-mediated food allergy. SignificanceAllergies are one of the most studied topics in the field of public health and novel protein allergens are found each year. Discovery of new principal and regional allergens has remarkable repercussions in precise molecular diagnostics, prognostics, and more specific immunotherapies. In this context, specific IgE is widely known to mediate physiopathology; however, allergies whose mechanism does not involve this immunoglobulin are poorly understood although their incidence has increased. Therefore, accurate diagnosis and adequate treatment are delayed with significant consequences on the health of pediatric patients. The study of type and subtypes of immunoglobulins associated with the immunoreactivity of cow's milk proteins together with an immunoproteomics approach allows better comprehension of physiopathology, brings the opportunity to discover new potential cow's milk protein allergens and may help in prognosis prediction (IgG1 occurrence as an early signal of possible risk toward development of IgE-mediated food allergy).

Malaria transmission-blocking conjugate vaccine in ALFQ adjuvant induces durable functional immune responses in rhesus macaques

Malaria transmission-blocking vaccines candidates based on Pfs25 and Pfs230 have advanced to clinical studies. Exoprotein A (EPA) conjugate of Pfs25 in Alhydrogel® developed functional immunity in humans, with limited durability. Pfs230 conjugated to EPA (Pfs230D1-EPA) with liposomal adjuvant AS01 is currently in clinical trials in Mali. Studies with these conjugates revealed that non-human primates are better than mice to recapitulate the human immunogenicity and functional activity. Here, we evaluated the effect of ALFQ, a liposomal adjuvant consisting of TLR4 agonist and QS21, on the immunogenicity of Pfs25-EPA and Pfs230D1-EPA in Rhesus macaques. Both conjugates generated strong antibody responses and functional activity after two vaccinations though activity declined rapidly. A third vaccination of Pfs230D1-EPA induced functional activity lasting at least 9 months. Antibody avidity increased with each vaccination and correlated strongly with functional activity. IgG subclass analysis showed induction of Th1 and Th2 subclass antibody levels that correlated with activity.

Variations in IgG antibody subclass responses to oral bacteria: Effects of periodontal disease and modifying factors.

Local and systemic IgG antibodies or oral bacteria have been described with periodontitis. We extended these observations by assessing the impact of a range of intrinsic factors on serum IgG subclass antibodies to both commensal and pathogenic oral bacteria that would contribute to variations in immune protection or disease susceptibility in periodontitis have not been described. Subjects (n=278) were classified as healthy, gingivitis, or periodontitis and categorized as mild, moderate, and severe periodontitis. Demographic stratification included sex, age, race/ethnicity, smoking, and obesity. Whole formalin-fixed bacteria were used as antigens to detect serum immunoglobulin (Ig)G subclass antibody levels using an ELISA. The greatest differences in variations in IgG subclasses occurred in periodontitis versus health or gingivitis to bacteria considered oral pathogens (eg, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, and Treponema denticola) with IgG1, IgG2, and IgG4 increased by three- to sevenfold with Pg. Differences in subclass levels and distribution were also observed related to disease severity, particularly related to individual subclass responses to Pg. Examination of the overall population showed that females had elevated antibody, reflected by elevated IgG2 amounts/proportions. The older group of subjects demonstrated elevated antibody to multiple oral bacteria, lacking any particular subclass pattern. IgG2 antibody to Aa and Pg was increased in smokers. Multiple IgG subclass antibody levels to oral pathogens were significantly decreased in the obese subset within this population. This investigation identified patterns of IgG subclass antibody responses to oral bacteria and demonstrated substantial effects of disease impacting the level and subclass distribution of antibody to an array of oral bacteria. Altered subclass antibody profiles most often in IgG2 levels and for antibody to P. gingivalis were found related to sex, age, disease severity, race/ethnicity, smoking, and obesity to both pathogens and commensal bacteria.

THE EFFECT OF HELMINTH CO-INFECTION ON MALARIA-SPECIFIC IMMUNOGLOBULIN G RESPONSES

BackgroundMalaria and helminthiases overlap extensively in their epidemiological distributions, and co-infections are common. Helminth infection has a profound effect on the immune system such as the induction of immuno-regulatory mechanisms...

Nanoparticle formulation enhanced protective immunity provoked by PYGPI8p-transamidase related protein (PyTAM) DNA vaccine in Plasmodium yoelii malaria model

We have previously reported the new formulation of polyethylimine (PEI) with gamma polyglutamic acid (γ-PGA) nanoparticle (NP) to have provided Plasmodium yoelii merozoite surface protein-1 (PyMSP-1) plasmid DNA vaccine with enhanced protective cellular and humoral immunity in the lethal mouse malaria model. PyGPI8p-transamidase-related protein (PyTAM) was selected as a possible candidate vaccine antigen by using DNA vaccination screening from 29 GPI anchor and signal sequence motif positive genes picked up using web-based bioinformatics tools; though the observed protection was not complete. Here, we observed augmented protective effect of PyTAM DNA vaccine by using PEI and γ-PGA complex as delivery system. NP-coated PyTAM plasmid DNA immunized mice showed a significant survival rate from lethal P. yoelii challenge infection compared with naked PyTAM plasmid or with NP-coated empty plasmid DNA group. Antigen-specific IgG1 and IgG2b subclass antibody levels, proportion of CD4 and CD8T cells producing IFN-γ in the splenocytes and IL-4, IFN-γ, IL-12 and TNF-α levels in the sera and in the supernatants from ex vivo splenocytes culture were all enhanced by the NP-coated PyTAM DNA vaccine. These data indicates that NP augments PyTAM protective immune response, and this enhancement was associated with increased DC activation and concomitant IL-12 production.

Association of HCMV specific IgG subclass antibody levels with gender and age

Infection with human cytomegalovirus (HCMV), a ubiquitously distributed herpesvirus, has been associated with human immune system aging. Especially total HCMV specific IgG antibody (AB) titres have been correlated with human frailty and mortality. The aim of this study was to further assess to which extent HCMV specific subclass AB titres differ between individuals and whether this is associated with age and gender.ELISA tests detecting HCMV specific IgG subclass 1 and 3 ABs, the main subclass ABs directed against HCMV, were developed. Using these ELISAs, 145 (73 female and 72 male) age-matched patients, aged 17 to 87years, were tested.The results showed that the IgG1 and 3 subclass distribution was highly variable among individuals, and that female persons had significantly higher IgG3 titres than age-matched male persons (p=0.0073). A correlation of the IgG3 subclass titres with age was found in female persons (Spearman r=0.305, p=0.0088).Thus the total HCMV specific IgG level appears to consist of a variable and gender associated distribution of IgG subclass ABs developed against persistent HCMV infection. The analysis of IgG subclass titres therefore could further be used to refine previously established clinical associations of aging with total HCMV specific IgG AB titres.

Curcumin-arteether combination therapy of Plasmodium berghei-infected mice prevents recrudescence through immunomodulation.

Earlier studies in this laboratory have shown the potential of artemisinin-curcumin combination therapy in experimental malaria. In a parasite recrudescence model in mice infected with Plasmodium berghei (ANKA), a single dose of alpha,beta-arteether (ART) with three oral doses of curcumin prevented recrudescence, providing almost 95% protection. The parasites were completely cleared in blood with ART-alone (AE) or ART+curcumin (AC) treatments in the short-term, although the clearance was faster in the latter case involving increased ROS generation. But, parasites in liver and spleen were not cleared in AE or AC treatments, perhaps, serving as a reservoir for recrudescence. Parasitemia in blood reached up to 60% in AE-treated mice during the recrudescence phase, leading to death of animals. A transient increase of up to 2–3% parasitemia was observed in AC-treatment, leading to protection and reversal of splenomegaly. A striking increase in spleen mRNA levels for TLR2, IL-10 and IgG-subclass antibodies but a decrease in those for INFγ and IL-12 was observed in AC-treatment. There was a striking increase in IL-10 and IgG subclass antibody levels but a decrease in INFγ levels in sera leading to protection against recrudescence. AC-treatment failed to protect against recrudescence in TLR2−/− and IL-10−/− animals. IL-10 injection to AE-treated wild type mice and AC-treated TLR2−/− mice was able to prolong survival. Blood from the recrudescence phase in AE-treatment, but not from AC-treatment, was able to reinfect and kill naïve animals. Sera from the recrudescence phase of AC-treated animals reacted with several parasite proteins compared to that from AE-treated animals. It is proposed that activation of TLR2-mediated innate immune response leading to enhanced IL-10 production and generation of anti-parasite antibodies contribute to protective immunity in AC-treated mice. These results indicate a potential for curcumin-based combination therapy to be tested for prevention of recrudescence in falciparum and relapse in vivax malaria.

Immunoglobulin G subclass antibody profiles in Porphyromonas gingivalis‐associated aggressive and chronic periodontitis patients

The immunoglobulin G (IgG) antibody response is considered to be protective and beneficial for the control of periodontal lesions. This study analysed IgG subclass antibody levels of Porphyromonas gingivalis in patients with both aggressive periodontitis (AgP) and chronic periodontitis (CP). Subgingival plaque and peripheral blood samples were collected from patients with localized AgP (n = 13), generalized AgP (n = 28) and generalized CP (n = 27) and from 14 periodontally healthy controls. P. gingivalis was identified in subgingival pockets using a polymerase chain reaction. Simultaneously, serum IgG subclass antibody against P. gingivalis whole cells/P. gingivalis fimbriae were measured using enzyme-linked immunosorbent assay. P. gingivalis was frequently detected in periodontitis patients. Anti-P. gingivalis whole cell IgG1 was elevated in all P. gingivalis-positive patients in the three periodontitis groups. Although increased anti-P. gingivalis IgG1 was also observed in the bacterium-positive healthy controls, the level was lower than that found in the three periodontitis groups. Levels of IgG1, IgG2, IgG3 and IgG4 to P. gingivalis did not differ among bacterium-positive patients in the three periodontitis groups; a significant increase of IgG2 level was not observed in localized AgP. Anti-fimbriae IgG subclass levels of IgG1, IgG2 and IgG4 did not differ among bacterium-positive subjects in all groups, while the anti-fimbriae IgG3 level in generalized CP was significantly higher than that in localized and generalized AgP. P. gingivalis infection elicited an IgG subclass antibody response in both periodontitis patients and healthy subjects, while higher anti-P. gingivalis IgG1 levels were found in the three periodontitis groups compared with the healthy control group.

Specific IgA subclass responses in serum and saliva: a 12-month follow-up study after parenteral booster immunization with tetanus toxoid

Specific IgA subclass antibodies against tetanus toxoid in serum, parotid saliva, and whole saliva were quantified after booster immunization. Samples from 14 healthy individuals were collected before and 1, 6, and 12 months after subcutaneous injection with Duplex ® (0.25 ml tetanus toxoid 30 Lf/mL and diphtheria 7.5 Lf/mL). Samples of whole saliva were also collected after 2 weeks. Specific IgA1 and IgA2 subclass antibodies to tetanus toxoid were quantified by enzyme-linked immunosorbent assay (ELISA). In this quantitative method, chimeric IgA1 and IgA2 antibodies directed against NP (4-hydroxy-3-nitrophenacetyl) were used as standards. Total levels of IgA1 and IgA2 were measured using a nephalometer or ELISA. Immunization with tetanus toxoid resulted in raised mean values of specific IgA1 and IgA2 antibodies against tetanus toxoid in serum after 1 month. Compared with the baseline, the mean value of specific IgA1 antibodies showed a 2.6-fold increase (mean value 10.47 µg/mL) in serum, and that of specific IgA2 antibodies a 2.7-fold increase (mean value 0.93 µg/mL). Specific IgA subclass antibody levels in parotid and whole saliva were unchanged after 1 month. The ratio of specific IgA subclass antibodies to total IgA subclass antibodies was 3 to 10 times higher in parotid saliva compared with whole saliva. In conclusion, subcutaneous booster immunization with tetanus toxoid induced immune responses of both antigen-specific IgA1 and IgA2 subclass antibodies in serum with the same increase, whereas the levels of specific IgA subclass antibodies in secretory fluids were unchanged. The ratio of specific IgA subclass antibodies to immunoglobulins was higher in parotid saliva compared with whole saliva.

Association between the IgG subclass response, inflammation and disease status in Helicobacter pylori infection.

In many viral, bacterial and parasitic infections the Immunoglobulin G (IgG) subclass response has been shown to correlate with severity of inflammation and disease outcome. The aim of the present study was to investigate the association between the IgG subclass response to Helicobacter pylori infection and disease and inflammation. Eighty-three symptomatic patients undergoing endoscopic examination were included in the study. Upon endoscopic examination, the presence of ulceration was noted and biopsy specimens were collected from the gastric antrum, body and transitional zone. Blood was also collected from each patient. Gastric biopsy sections were graded using the Sydney system. H. pylori specific IgG, IgG1, IgG2, IgG3 and IgG4 were measured by ELISA. The IgG subclass was also examined retrospectively in sera collected from 20 patients previously proven to have duodenal ulcer (DU). The results of histological examination and IgG serology showed 35 subjects to be H. pylori negative and 48 to be H. pylori positive. Of the 48 H. pylori positive subjects, 25 were diagnosed with functional dyspepsia (FD), 14 with current DU and 9 with evidence of past DU. Significantly higher levels of IgG2 antibodies were found in patients with DU as compared with patients with FD (P < 0.01). In addition, significantly higher IgG3 subclass antibody levels were associated with chronic inflammatory cells in the body (P < 0.05) and active inflammatory cells in the transitional zone (P < 0.01). A significantly increased level of IgG1 antibodies was associated with lower levels of colonization in the gastric antrum. The results of this study suggest that the IgG subclass response in subjects infected with H. pylori may be a marker of DU disease as well as increased levels of inflammation.

Association Between the IgG Subclass Response, Inflammation and Disease Status in Helicobacter pylori Infection

Background: In many viral, bacterial and parasitic infections the Immunoglobulin G (IgG) subclass response has been shown to correlate with severity of inflammation and disease outcome. The aim of the present study was to investigate the association between the IgG subclass response to Helicobacter pylori infection and disease and inflammation. Methods: Eighty-three symptomatic patients undergoing endoscopic examination were included in the study. Upon endoscopic examination, the presence of ulceration was noted and biopsy specimens were collected from the gastric antrum, body and transitional zone. Blood was also collected from each patient. Gastric biopsy sections were graded using the Sydney system. H. pylori specific IgG, IgG1, IgG2, IgG3 and IgG4 were measured by ELISA. The IgG subclass was also examined retrospectively in sera collected from 20 patients previously proven to have duodenal ulcer (DU). Results: The results of histological examination and IgG serology showed 35 subjects to be H. pylori negative and 48 to be H. pylori positive. Of the 48 H. pylori positive subjects, 25 were diagnosed with functional dyspepsia (FD), 14 with current DU and 9 with evidence of past DU. Significantly higher levels of IgG2 antibodies were found in patients with DU as compared with patients with FD (P < 0.01). In addition, significantly higher IgG3 subclass antibody levels were associated with chronic inflammatory cells in the body (P < 0.05) and active inflammatory cells in the transitional zone (P < 0.01). A significantly increased level of IgG1 antibodies was associated with lower levels of colonization in the gastric antrum. Conclusion: The results of this study suggest that the IgG subclass response in subjects infected with H. pylori may be a marker of DU disease as well as increased levels of inflammation.

Cord blood levels of immunoglobulin G subclass antibodies to food and inhalant allergens in relation to maternal atopy and the development of atopic disease during the first 8 years of life.

Factors that either protect from or enhance the development of atopic disease appear to be acting early in life. The gestational environment, including maternal immune responses, such as transplacentally transferred immunoglobulin (Ig) G antibodies to allergens, may be of importance in this respect, since allergen-specific immunity has been demonstrated to develop in utero. To evaluate the relation between cord blood IgG subclass antibodies to allergens, maternal atopy and development of atopic disease in the children. The study group comprised a cohort of 96 children participating in a prospective study up to 8 years of age. Cord blood IgG subclass antibodies to ovalbumin, beta-lactoglobulin, Bet v 1 and cat dander were analysed by ELISA. The levels of all IgG subclass antibodies to ovalbumin and rBet v 1 were higher in newborn infants with an atopic mother, as compared with babies with nonatopic mothers. IgG1 antibody levels to cat and IgG4 antibody levels to beta-lactoglobulin and cat were also higher in atopic than in nonatopic mothers, whereas the other subclass antibody levels to those allergens were similar. High levels of cord blood IgG antibodies to cat and birch, but not to the food allergens, were associated with less atopic symptoms in the children during the first 8 years of life. Moreover, children who developed IgE antibodies to cat had lower levels of IgG antibodies to that allergen at birth. High levels of cord blood IgG subclass, especially IgG4, antibodies to food and inhalant allergens are associated with maternal atopy. High levels of IgG antibodies to inhalant, but not food, allergens are associated with less development of atopy in the children.

Exaggerated systemic antibody response to mucosal Helicobacter pylori infection in IgA nephropathy

Numerous studies in the literature report aberrant immune responsiveness in immunoglobulin A (IgA) nephropathy. However, all these studies investigate immune responses invoked by an artificially engineered antigen challenge. For the first time in IgA nephropathy, we report the systemic humoral responses generated as part of an active mucosal immune response against a common environmental pathogen, Helicobacter pylori (Hp). We studied 22 patients with IgA nephropathy and 9 controls without renal disease who were shown to be infected with Hp, using a 13C-urea breath test. Hp antigen-specific enzyme-linked immunosorbent assays were established to measure the anti-Hp IgA, IgG, and IgA and IgG subclass antibody levels. In addition, anti-Hp responses in the monomeric and polymeric (pIgA) fractions of serum IgA were measured after separation by gel filtration high-performance liquid chromatography. IgA nephropathy was associated with both a greater rate of IgA anti-Hp seropositivity (P < 0.05) and a more pronounced IgA anti-Hp antibody response (P < 0.01). In almost all cases, IgA anti-Hp was IgA1, and more than 90% was polymeric. There was no difference in the frequency of IgG anti-Hp seropositivity, but patients produced a much greater IgG anti-Hp response (P < 0.01). In addition, the IgG subclass responses were markedly different, with IgG1 predominant in controls and IgG2 and IgG3 the major subclasses produced in IgA nephropathy. We have shown an exaggerated systemic antibody response to mucosal infection caused by Hp in patients with IgA nephropathy, predominantly consisting of pIgA1, IgG2, and IgG3. This suggests that in IgA nephropathy, not only is pIgA1 production poorly controlled, but regulation of IgG isotype switching in response to mucosal pathogens is also deranged.

Human IgG subclass responses in relation to serum bactericidal and opsonic activities after immunization with three doses of the Norwegian serogroup B meningococcal outer membrane vesicle vaccine

Ten adult volunteers, with low prevaccination levels of serum IgG antibodies against meningococcal antigens (<1 μg ml −1), received three doses of the Norwegian group B meningococcal outer membrane vesicle (OMV) vaccine intramuscularly at weeks 0, 6 and 46. Anti-OMV IgG subclass responses were measured and compared with serum bactericidal activity (SBA) and opsonic activity against the vaccine strain 44/76. All vaccinees showed an IgG1 antibody response after each vaccine dose. The vaccine-induced median serum IgG1 antibody levels were 16, 17 and 18 μg ml −1 2–6 weeks after the first, second and third dose, respectively. Three vaccinees showed a weak IgG3 response after the first dose, whereas 8 and 9 showed a response after the second (median=10 μg ml −1) and third dose (median=10 μg ml −1), respectively. Low levels of anti-OMV IgG2 antibodies were found, whilst specific IgG4 antibodies were only detected for one vaccinee. The vaccine induced at least a fourfold increase in SBA titre in 8 vaccinees after the first dose, in 9 vaccinees after 2 doses and in all vaccinees after 3 doses. A positive correlation was found between IgG1 subclass antibody levels and SBA ( r=0.62, P<0.0001). Elevated opsonophagocytic activity, measured as respiratory burst (RB), was observed in all vaccinees after one vaccine dose and usually increased after 2 and 3 doses. A strong positive correlation was found between IgG1 antibody levels and RB ( r=0.76, P<0.0001). In conclusion, we have shown that systemic meningococcal OMV vaccination in adult vaccinees mainly induced IgG1 antibodies which correlated with bactericidal and opsonic activity, but also a considerable amount of IgG3 antibodies, which, in contrast to the IgG1 response, was induced only after 2 or 3 vaccine doses and declined more rapidly.

Specific IgG subclass antibody pattern toAspergillus fumigatus in patients with cystic fibrosis with allergic bronchopulmonary aspergillosis (ABPA)

BACKGROUNDIgG and IgG subclass antibodies to Aspergillus fumigatus (A fumigatus) were measured in a large population of patients with cystic fibrosis to elucidate a putative antibody pattern specific for allergic...

Exposure to cow’s milk during the first 3 months of life is associated with increased levels of IgG subclass antibodies to β-lactoglobulin to 8 years

Background: Exposure to allergens early in life influences the development of allergen-specific immune responses. In animal models, the development of tolerance to proteins delivered to the gastrointestinal and the respiratory mucosa is influenced by age and genetic background. Late introduction of cow’s milk in infants is associated with slower increase and lower peak IgG antibody responses to milk during early childhood, but the long-term effects have not been investigated, nor is the relation to atopic disease later in life clear. Objective: The purpose of this study was to investigate the development of IgG subclass antibodies to β-lactoglobulin in relation to early exposure to cow’s milk, atopic heredity, and the development of atopic disease. Methods: IgG subclass antibodies to β-lactoglobulin were analyzed by ELISA at birth, at 6 and 18 months, and at 8 years in 96 children followed prospectively. Results: The levels of IgG subclass antibodies to β-lactoglobulin peaked in early childhood and then declined up to 8 years of age. Exposure to cow’s milk during the first 3 months of life was associated with high IgG subclass antibody levels to β-lactoglobulin up to 8 years, particularly in children with maternal atopy. Children with atopic symptoms and sensitivity to allergens often had high levels of IgG 4 antibodies to β-lactoglobulin at 8 years of age, even if they were not exposed to cow’s milk during the first 3 months of life. Furthermore, atopic dermatitis was associated with high levels of IgG subclass antibodies to β-lactoglobulin in early childhood. Conclusions: IgG subclass antibody levels to milk peak during early infancy, with particularly high levels in children with atopic dermatitis, and decline thereafter. Exposure to cow’s milk during early infancy has long-lasting effects on the humoral antigen-specific responses, indicating less effective tolerance-inducing mechanisms in the intestinal mucosa during the first months of life. (J Allergy Clin Immunol 1998;102:671-8.)

Rapid evaluation of serum and gingival crevicular fluid immunoglobulin G subclass antibody levels in patients with early-onset periodontitis using checkerboard immunoblotting.

A method was developed to evaluate the presence of immunoglobulin G (IgG) subclass (1-4) antibody to Actinobacillus actinomycetemcomitans, serotype b (strain Y4) in patients with early-onset periodontitis on a single nitrocellulose membrane. Sera from 30 early-onset periodontitis patients and gingival crevicular fluid samples from 2 patients were collected and tested with four different preparations of A. actinomycetemcomitans (Y4). The principle steps of the assay are: a) binding of the bacterial antigen (Y4) and the anti-human IgG antibody (capture antibody) in parallel lanes on nitrocellulose membranes; b) incubation of known concentrations of the IgG subclasses 1, 2, 3 and 4, as well as a dilution of serum and/or gingival crevicular fluid from patients in lanes perpendicular to the antigen lanes; c) incubation of the membranes with the corresponding peroxidase conjugated anti-human IgG subclass secondary antibody; d) detection of positive signals by enhanced chemiluminescence. The blots were evaluated by visual comparison to a series of blots containing known concentrations of IgG subclasses. The method was used to rapidly screen a relatively large number of patient sera and gingival crevicular fluid samples for IgG subclasses in a cost-effective assay. The predominant IgG subclass found in early-onset periodontitis was IgG2.

IgG1 and IgG4 serum antibody responses in asymptomatic and clinically expressed cystic echinococcosis patients

IgG1 and IgG4 subclass antibody responses were investigated in two clinically different groups of hepatic cystic echinococcosis (CE) patients. One group consisted of surgically proven CE cases (clinically expressed hospitalized cases) and a second group comprised asymptomatic CE patients (first time community detected cases) diagnosed by portable ultrasound and serology in four different endemic communities. Fifty eight sera from surgically proven CE patients and 133 sera from asymptomatic ultrasound diagnosed CE cases were screened by ELISA using purified human hydatid cyst fluid antigen B for total specific IgG, IgG1 and IgG4 antibodies. Fifty sera from healthy individuals from within endemic regions were used as control negatives. Compared to control negatives total IgG antibody levels were elevated in both surgically proven (85%) and asymptomatic CE cases (77%) but there was no significant difference between the two groups. IgG1 subclass antibody levels were also elevated in both surgically proven (55%) and asymptomatic cases (58%) compared to endemic controls and similarly the difference in this response was not significant between these two groups. In contrast the prevalence of IgG4 antibodies in surgically confirmed chronic CE patients was greater (71%) than the respective IgG1 antibody levels (55%). The greatest difference in specific antibody response, between advanced surgically confirmed CE patients and ultrasound detected asymptomatic CE cases, was observed for IgG4 antibody levels which were detected in 71% of symptomatic compared to only 23% of asymptomatic patients ( P<0.0001). These observations confirm that IgG4 is an important diagnostic parameter for clinically expressed cystic echinococcosis in humans and suggest that a switch from a predominant IgG1 response to IgG4 might occur in CE patients as the disease progresses.

Significantly increased IgG2 subclass antibody levels to Blastocystis hominis in patients with irritable bowel syndrome.

Blastocystis hominis is a common intestinal parasite of humans in the tropics whose pathogenic role is in dispute. Its presence has been reported in a variety of intestinal disorders resembling irritable bowel syndrome (IBS) such as diarrhea, anorexia, and flatulence. We have therefore investigated a possible link between IBS and blastocystosis by determining IgG antibody levels to B. hominis in patients with IBS. Levels of IgG antibodies were significantly elevated in patients with IBS compared with asymptomatic controls (P < 0.0001, by Student's t-test) in both B. hominis stool culture-positive and stool culture-negative IBS patients. When IgG antibodies were divided into their respective subclasses, only IgG2 levels were significantly increased in IBS patients compared with asymptomatic controls, indicating that the predominant response in these patients may be directed to carbohydrate antigens. The diagnostic usefulness of this test in IBS patients remains to be established because these data are only suggestive of a possible link between B. hominis and IBS. However, we hope that this antibody test will help in elucidating the controversy that surrounds the role of B. hominis as a pathogen at present.

The Effects of Polypodium leucotomos Extract on the Specific Antibody Production Patterns in BALB/c Mice Immunized with Third Stage Larvae Antigens of Anisakis simplex

The immunomodulant effects of Polypodium leucotomos extracts have been tested in BALB/c mice immunized with the third larval stage antigens of Anisakis simplex. A decrease in the specific antibodies was observed in animals treated either before or after immunization. This decrease was mainly in the IgG 1 isotype. In animals treated before immunization the reduction of the IgG 1 subclass was dramatic in relation to the untreated group while the IgG 2b subclass appeared earlier and at slightly higher levels than in the control group. In animals treated after immunization a reduction of the IgG 1 subclass was also observed although not as marked as in the preimmunization group. IgG 2a sub-class antibody levels were higher than those of the untreated group. As in the preimmunization group, the IgG 2b subclass antibodies appeared earlier but they remained at similar levels to those observed in the control group.