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The role of the cytokines interleukin-4 and interferon-gamma in the regulation of IgE responses in the mouse and man have focused on the role of CD4 T cells. In the rat, antigen-specific CD8 T cells, generated following inhalation of antigen, have been shown to be capable of suppressing IgE responses. Repeated intraperitoneal injections of 1 ng ricin and 1 microgram antigen established a long-lived IgE response in both low- and high-IgE responder rat strains (Wistar and Brown Norway). The duration of the IgE antibody response was 204 and 248 days, respectively. Total IgE levels rose from 30 +/- 20 to 39,000 +/- 7,500 ng/ml in the Wistar rat and from 120 +/- 100 to 47,000 +/- 8,000 ng/ml in the Brown Norway rat. An even greater (10(4)-fold) increase was seen in antigen-specific IgE antibody levels. Ricin alone had no effect and concomitant or prior stimulation with antigen was required. The proportion of CD4+ and CD8+ cells present in the spleen at the peak of the IgE response was markedly increased compared with animals given ricin or antigen alone. Furthermore, CD8 T cells were approximately 100 times more sensitive to ricin than CD4 T cells. These data suggest that enhancement of IgE responses in ricin-treated animals results from the selective deletion of T cells which suppress IgE and are of the CD8 phenotype.

A study was carried out on beekeepers and their families. 34 subjects with a history of bee sting allergy and also a positive radioallergosorbent test (RAST) to honey bee venom were compared with 47 subjects with neither. 16 subjects in the bee allergy group (47%) had a history of atopic diseases as compared to 6 (13%) in the control group (p less than 0.01). 15 in the bee allergy group (44%) had at least one positive RAST to three common inhalant allergens as compared to 5 (11%) in the control group (p less than 0.01). These results suggest that, although bee sting allergy frequently affects non-atopic subjects, sensitization among beekeepers and their families occurs more readily in atopics. It is notable that a large proportion of patients injected with foreign proteins (the control group) either never develop venom-specific IgE antibodies or have suppressed their production.

The response of 15 strongly bee-venom-allergic patients to highly purified venom allergens was compared using skin prick test titration, peripheral blood leukocyte (PBL) histamine release and radioallergosorbent test with three highly purified bee venom allergens: phospholipase (PLA2), hyaluronidase (HYAL) and acid phosphatase (ACID P). Sensitivity to the three allergens ranked in the same order for all three tests and in each case PLA2 was found to the most potent allergen. In the presence of hyperimmune beekeeper plasma, maximum histamine release was reduced significantly for all three allergens (p less than 0.001). Furthermore, hyperimmune beekeeper plasma increased the amount of allergen required for a comparable release of histamine (mean shift in dilution curve PLA2 917-fold; HYAL, 492-fold; ACID P, 61-fold). The release of histamine from whole blood was also compared with PBL + 10% normal human serum (NHS). For all three allergens maximum release was much lower from whole blood compared with washed cells + 10% NHS (p less than 0.001). These data confirm PLA2 as the major bee venom allergen by all three tests. Hyperimmune beekeeper plasma reduces maximum histamine release and increases its threshold. Histamine release in response to ACID P appears harder to block with hyperimmune beekeeper plasma than that provoked by PLA2 or HYAL (p less than 0.01). Whole blood releases less histamine and requires more allergen than washed cells, indicating that sensitivity of PBL in vivo is unlikely to be as great as washed PBL in vitro.

We have measured in allergic and non-allergic beekeepers IgE and IgG antibodies to the bee venom allergens, phospholipase A2 (PLA2) and hyaluronidase (HYAL), by the radioallergosorbent test (RAST) and a 125I-radiolabelled antigen-binding assay. The absolute amount of IgG antibody in a reference serum was determined by saturation analysis using 125I-radiolabelled PLA2 and HYAL. Using monoclonal anti-IgE coated microtitre plates, the absolute amount of IgE antibody to the same antigen was also determined by saturation analysis. Regardless of the IgE response to the different allergens, IgG antibody concentrations to PLA2 were invariably higher than those to HYAL. In addition, the ratio of IgG to IgE antibody was higher for PLA2 (220:1) than for HYAL (10:1). Higher levels of IgG antibody to both allergens (especially HYAL) were found in those who had had prolonged exposure to bee stings. These data suggest that the level of IgG antibody produced is related to the dose administered, while the amount of IgE antibody may be regulated by other factors.

The effect of the castor bean toxin, ricin, on rat spleen and mesenteric lymph node cell cultures was tested. Rat lymphocytes were cultured for 2 days in the presence of optimal doses of the mitogens phytohaemagglutinin, conconavalin A, and lipopolysaccharide. At concentrations of 10(-3) to 10(-5) mg/ml ricin, spontaneous and mitogen-driven proliferation were inhibited. At lower concentrations, spleen but not mesenteric lymph node cells showed enhanced proliferation. Studies are continuing to identify subpopulations of lymphocytes with increased or decreased sensitivity to ricin and their role in the regulation of IgE.

The antibody response to phospholipase A2 (PLA2) was studied by using radiolabelled antigen-binding techniques for both IgG and IgE antibodies. The results are compared with results for antigen P1 from Dermatophagoides pteronyssinus and for rye I from rye grass pollen. The response to PLA2 differs from the response to the inhalant allergens in several ways: almost all exposed bee keepers produce IgG antibodies to PLA2; among the allergic bee keepers there was no quantitative correlation between IgG and IgE antibodies; and the antibodies to PLA2 are not produced locally in the nose. Bee keepers with hay fever did not appear to have an increased incidence of allergic reactions to bee venom. In addition, the bee keepers with hay fever and the same levels of IgG and IgE antibodies to rye I as other individuals with hay fever. However, the bee keepers who had no history of hay fever were found to have high levels of IgG antibodies to rye I. These antibodies did not cross-react with PLA2 or bee venom, and the results suggest that bee stings may act as an adjuvant for IgG antibodies to pollen antigens.

IgG and IgE antibody levels have been followed for a period of 2 years in patients receiving immunotherapy with bee and wasp venom. 106 adult patients who had had anaphylactic reactions to wasp stings had initially low IgG antibody levels to wasp venom which rose with therapy (p less than 0.001). IgE antibody levels also showed an initial rise but subsequently fell (p less than 0.001). The pattern was similar to that previously reported in children who had had anaphylactic reactions to bee stings, but who, after a course of immunotherapy, were able to tolerate stings with impunity. 60 adults who had had anaphylactic reactions to bee stings showed a different pattern, with initially high IgG antibody levels which did not rise further. Since two thirds of this group were beekeepers or members of beekeeper's families, the high initial IgG antibody levels could have been a response to the frequent stings to which such individuals are prone. The fact that high levels did not protect against anaphylaxis shows, however, that the classical concept of 'blocking antibody' is in need of revision.

We studied 55 subjects who had had anaphylactic reactions to bee stings within the previous 3 years. 38 out of 54 tested had IgE antibody to honey bee venom (HBV) as measured by radioallergosorbent test (RAST). On skin testing, 30 out of 34 had a positive test to HBV. Of these, 26 had a positive RAST. A positive skin test to HBV at high dilution or else a high anti-HBV RAST score appeared to identify those who, in a 6-month follow-up period, were at risk of developing further anaphylaxis following bee stings or immunotherapy. Of the two tests, RAST appeared to be the less sensitive. Measurements of IgG antibody to phospholipase A were seldom available for the period immediately preceding an anaphylactic episode and proved to be a poor means of predicting the liability to bee sting anaphylaxis in subsequent months.

The relationship between serum IgE and castor bean (Ricinus communis)-specific IgE was studied in a group of dockworkers and other residents of Marseilles who were diagnosed as having castor bean allergy. The diagnosis was made because they had asthma or other allergic symptoms together with a positive skin test to castor bean. Total serum IgE was found to be higher in castor-bean-allergic patients (mean = 174 IU/ml) than in a control group of local blood donors (mean = 66 IU/ml). IgE levels were not as high as those found in the sera of a previously identified group of castor-bean-allergic dockworkers from Port Sudan (mean = 902 IU/ml). Castor-bean-specific IgE was demonstrated in the serum of 91% of the allergic group and there was some correlation with total IgE (r = 0.53, p less than 0.01). The proportion of IgE antibody specific for castor bean was determined in 20 castor-bean-sensitive patients using the radioallergosorbent test and was found to vary between 9 and 64% (mean = 38% or 265 ng/ml). No castor-bean-specific IgE antibody was detected in the control group.

Indirect ELISA is widely used to detect specific antibodies but can suffer from high non-specific binding-particularly of IgG. The use of affinity-purified rabbit antibody-coated microtitre plates to bind antigen greatly increases sensitivity without a significant increase in non-specific binding of IgG. Capture, competitive and sandwich assay procedures gave comparable results for IgG antibodies; but only the sandwich assay was suitable for detection of IgE antibodies.

We have measured IgG1, 2, 3 and 4 subclass antibodies to gliadin and casein in normal controls, treated and untreated coeliac patients. The amount of antibody in each subclass was determined separately as a titre and as a concentration, by reference to a standard curve. Both methods of quantitation, carried out in separate assays, yielded comparable results (IgG antibody r = 0.86, IgG1 antibody r = 0.89). In all the coeliac patients, the majority of antibody was IgG1, and little IgG2, 3, 4 antibody was detected. Untreated coeliac patients had higher titres of IgG and IgG1 antibodies to gliadin and casein compared with normal controls. In the treated coeliac patients compared with the normal controls, there was no significant difference between the titres of IgG and IgG1 antibodies to gliadin but there were higher titres to casein.