Current practice of allergy diagnosis and the potential impact of regulation in Europe.

In the European Union (EU), the regulatory framework regarding diagnostic allergen extracts is currently in the process of being implemented at the national level. Due to these regulations, the initial and periodic renewal expenses for the registration of diagnostic allergen extracts may render extract production unprofitable. Consequently, many extracts may be at risk of removal from the market. The current survey, which was conducted by a task force of the European Academy of Allergy and Clinical Immunology, aimed to assess the current practice of allergy diagnosis in Europe. This survey revealed that skin tests continue to be the main diagnostic procedure and are used as the first option in almost two-third of all types of allergic diseases and in 90% of individuals suffering from respiratory allergies. Therefore, there is a need to ensure the availability of high-quality allergen extracts to maintain the common diagnostic procedures used by EU professionals. To reach this goal, it is necessary to align efforts and establish active partnerships between manufacturers, relevant scientific societies, consumer organizations and authorities to maintain the availability of these diagnostic tools.

Immunotherapy is allergen-specific: a double-blind trial of mite or timothy extract in mite and grass dual-allergic patients.

BACKGROUND: One hundred years ago, Noon [Lancet 1911;1:1572-1573], using conjunctival provocation testing (CPT), was the first to demonstrate the effectiveness of subcutaneous immunotherapy (SCIT) in grass-allergic subjects with hay fever. In this centenary year, we present data that, by use of CPT and allergen-specific IgG, replicate this observation and additionally confirm the allergen specificity of SCIT by using a double-blind design employing either grass or mite SCIT in dual grass- and mite-allergic individuals. METHODS: Twenty adults (11 females) with perennial rhinoconjunctivitis and exacerbation of symptoms during the grass pollen season and in the autumn had immediate skin and conjunctival sensitivity and raised specific IgE to both Dermatophagoides farinae and Phleum pratense. Participants were randomly assigned to either timothy or D. farinae immunotherapy for 3 years. CPT and specific IgG tests to both allergens were performed annually. After 3 years, subjects gave their blinded overall evaluation. RESULTS: Six mild-to-moderate general reactions occurred in 2 timothy- and 4 mite-treated patients. Four of these patients and 2 other patients withdrew from the study. Seven patients in each group completed the study. After 3 years of immunotherapy, the timothy CPT threshold concentration had increased 16- fold in timothy-treated patients (p < 0.05; between-group change, p < 0.05). The increase in the mite CPT threshold in mite- compared to grass-treated patients was 31-fold (p < 0.05). The overall assessment of conjunctival sensitivity was highly significant in favour of treatment (p < 0.015), as was that of allergen-specific IgG (p < 0.0001). CONCLUSIONS: Allergen immunotherapy is allergen species-specific, as judged by decreased conjunctival sensitivity and changes in allergen-specific IgG concentrations.

Immunotherapy with monomethoxypolyethylene glycol modified allergens.

On the basis of the results of experiments in naive mice, i.e., in mice which had not been presensitized, it was anticipated that mPEG-modified allergens would suppress the specific IgE response in allergic humans. However, only minor or no suppression of IgE antibodies was induced on administration of mPEG conjugates in allergic patients with a longstanding IgE response. This observation was later confirmed in pre-sensitized animals. The mPEG-modified allergens can be synthesized in a reproducible manner. This makes mPEG-modified allergens suitable for production and quality control. There is indirect evidence, i.e., identification of IgE and IgG antibody stimulation against all identified allergens, that the relevant determinants are available for processing by the immune system. By contrast, the polymerization of allergens by formaldehyde or glutaraldehyde leads to ill-defined, cross-linked high molecular weight materials. As in the case of other modified allergens, mPEG-modified allergens have been developed primarily because of their lower allergenicity than the original allergen preparations. However, they also retain clinical efficacy of the same magnitude as that induced by unmodified allergen preparations, which is a prerequisite for clinical use. IT with mPEG-modified pollen allergen preparations has been proven to be as effective as IT with unmodified allergens, when these preparations are given in similar doses. Furthermore, a more pronounced beneficial effect can be obtained with higher doses of mPEG-modified allergen, without the risk of side effects which are often precipitated by unmodified allergens. Therefore, it seems reasonable to suggest that children and young adults with developing sensitivity to perennial allergens, and without chronic changes of the bronchial mucosa, are the most suitable candidates for IT with mPEG-modified allergen preparations. Mite asthma, which always includes bronchial inflammation and nonspecific hyperresponsiveness, represents a more complicated model. In mite asthmatics mPEG-modified mite allergen preparations were safer than the corresponding unmodified preparation. Although bronchial sensitivity to allergen and histamine did not change significantly in mite asthmatics during the observation time, it appears that IT with mPEG-modified mite allergen led to a decrease in the releasability of histamine from skin mast cells, as measured by SPT, to a similar degree as observed in pollinosis patients. Therefore, it may be inferred that the allergic component of asthma might be influenced by IT with mPEG allergens. As a consequence, before the start of IT, the degree of reversibility of pathophysiological changes in the bronchial wall should be considered.(ABSTRACT TRUNCATED AT 400 WORDS)

Assay of the beta-glucosidase activity with natural labelled and artificial substrates in leukocytes from homozygotes and heterozygotes with the Norrbottnian type (Type 3) of Gaucher disease.

Leukocytes were isolated from 14 patients (7 males and 7 females ) with Gaucher disease of the Norrbottnian type (Type 3), 32 obligate heterozygotes (16 males and 16 females) for this disease and 20 controls (10 males and 10 females). After collection, the cells were transported in dry ice to the laboratory, where they were assayed. The assays were repeated after the cells had been stored for 12 months. beta-Glucosidase activity was assayed with D-[glucose-U-14C]glucosylceramide at pH 5.8 with Cutscum-Na-cholate as a detergent and 4-methylumbelliferyl-beta-glucoside at pH 4.1 with Triton-Na-taurocholate as a detergent. The activities of two marker enzymes, 4-methylumbelliferyl-beta-galactosidase and N-acetyl-beta-glucosaminidase, were assayed in aliquots of the same leukocyte samples. The activity of beta-galactosidase remained constant during storage, N-acetyl-beta-glucosaminidase increased, while beta-glucosidase decreased as assayed with the natural as well as with the artificial substrate. beta-Glucosidase activity was significantly lower in the female than in male controls and heterozygotes. When assayed with natural substrate beta-glucosidase activity in leukocytes from the male patients was 6--12% of the control mean value and 10--15% in those from the female patients. The corresponding figures found when the artificial substrate was used were 15--30% and 22--45%. The values for the heterozygotes were respectively 42--68% and 34--79% with the natural substrate, and 33--82% and 51--109% with the artificial substrate. No correlation was found between the age of the patient and the beta-glucosidase activity.

Gaucher disease (type III): intellectual profile.

In 20 patients with Gaucher disease type III (Norrbottnian variant), long-term intellectual prognoses were analyzed on the basis of psychometric tests which were performed on an average of five tests per patient. Intellectual delay was not found to be characteristic of the early stages of the disease. Slow regression occurred through childhood and adolescence. Patients splenectomized at an early stage averaged lower IQ scores in the long-term than those in whom the spleen had been spared as long as possible. These data added to other evidence of increased neurologic and other organ impairment after splenectomy, support the view that the spleen should not be removed in other than emergent situations.

Attempt at enzyme replacement by organ transplantation: renal transplantation in Gaucher disease.

Following renal transplantation, hepatic glucocerebroside deposits in a child with Gaucher disease were reduced. This suggests that enzyme replacement had been achieved.

Specific immunotherapy has long-term preventive effect of seasonal and perennial asthma: 10-year follow-up on the PAT study.

BACKGROUND: 3-year subcutaneous specific immunotherapy (SIT) in children with seasonal allergic rhinoconjunctivitis reduced the risk of developing asthma during treatment and 2 years after discontinuation of SIT (5-year follow-up) indicating long-term preventive effect of SIT. OBJECTIVE: We evaluated the long-term clinical effect and the preventive effect of developing asthma 7-years after termination of SIT. METHODS: One hundred and forty-seven subjects, aged 16-25 years with grass and/or birch pollen allergy was investigated 10 years after initiation of a 3-year course of SIT with standardized allergen extracts of grass and/or birch or no SIT respectively. Conjunctival provocations were performed outside the season and methacholine bronchial provocations were performed during the season and winter. Asthma was assessed by clinical evaluation. RESULTS: The significant improvements in rhinoconjunctivitis and conjunctival sensitivity persisted at the 10-year follow-up. Significantly less actively treated subjects had developed asthma at 10-year follow-up as evaluated by clinical symptoms [odds ratio 2.5 (1.1-5.9)]. Patients who developed asthma among controls were 24/53 and in the SIT group 16/64. The longitudinal treatment effect when adjusted for bronchial hyper-responsiveness and asthma status at baseline including all observations at 3, 5 and 10 years follow-up (children with or without asthma at baseline, n = 189; 511 observations) was statistically significant (P = 0.0075). The odds ratio for no-asthma was 4.6 95% CI (1.5-13.7) in favor of SIT. CONCLUSION: A 3-year course of SIT with standardized allergen extracts has shown long-term clinical effects and the potential of preventing development of asthma in children with allergic rhinoconjunctivitis up to 7 years after treatment. CLINICAL IMPLICATION: Specific immunotherapy has long-term clinical effects and the potential of preventing development of asthma in children with allergic rhino conjunctivitis up to 7 years after treatment termination.

Five-year follow-up on the PAT study: specific immunotherapy and long-term prevention of asthma in children.

BACKGROUND: A 3-year course of specific immunotherapy (SIT) in children with hay fever to grass and/or birch pollen significantly reduced the risk of developing asthma. To investigate the long-term preventive effect, we performed a follow up--2 years after termination of immunotherapy. METHODS: A total of 183 children, aged 6-14 years with grass and/or birch pollen allergy could be investigated 2 years after discontinuation of SIT or no treatment. Conjunctival provocation tests (CPTs) and methacholine bronchial provocation tests were carried out during the season and winter after 5 years. The development of asthma was assessed by clinical evaluation. RESULTS: The significant improvement in hay fever and CPT results observed after 3 years of SIT persisted at the 5-year follow-up. No difference in bronchial responsiveness to methacholine was found after 5 years because of spontaneous improvement during the follow-up period in the control patients. The immunotherapy-treated children had significantly less asthma after 5 years as evaluated by clinical symptoms [odds ratio 2.68 (1.3-5.7)] in favor of SIT for prevention of development of asthma and significantly less patients reported an increase in asthma scores (P < 0.01). CONCLUSION: Immunotherapy for 3 years with standardized allergen extracts of grass and/or birch shows long-term clinical effect and preventive effect on development of asthma in children with seasonal rhinoconjunctivitis.

Skin test in diagnosis of food allergy.

Skin tests have for many years been widely used in the diagnosis of allergic diseases. Standardized allergic tests are commonly available for the diagnosis of inhalant allergy. However food allergens have been studied and no standardized preparations are available. Methods for skin testing vary between regions. In North America, the intradermal skin-test method dominates. On the other hand, pediatricians and Europeans favor the use of skin-prick test, as it is less painful and gives the same information as the intradermal method. Methods for interpretation also vary between regions and groups of doctors. Furthermore, criteria for disease (i.e., presence of food allergy) also varies. Because of the lack of standardized allergen techniques and method, there are difficulties in understanding and using the new data presented in scientific journals. In this article I discuss these difficulties and propose how to handle these problems, concentrating on the skin-prick test method.

The batch-to-batch variation of the potency of dog- and cat-allergen-coated lancets. Evaluation by skin prick testing.

In clinical work I formed the impression that the potency of Phazet lancets, for cat and dog allergens, was lower than earlier and too low to diagnose all clinically sensitive children with cat and dog allergy. Therefore, I decided to investigate the potency of newly produced and several years old Phazet lancets coated with dog and cat allergens. Twenty-six adults with case histories of cat and/or dog allergy and skin reactive to extracts of either dog dander or cat epithelium were skin prick tested in duplicate with one old batch and one newly produced batch of Phazet dog and/or cat lancets as well as the in-house reference of cat and/or dog allergen, 100,000 BU/ml. The potency of Phazet lancets in relation to the standard was evaluated by the median slope of the allergen dose-response relationship. The potency of the old dog lancets (mean 48,900 BU/ml) was higher (NS) than that of the newly produced lancets (mean 28,000 BU/ml), and the potency of both types of lancets was significantly less (P < 0.001) than that of the dog allergen in-house standard. On the other hand, the potency of newly produced (123,000 BU/ml), but not old (108,000), cat-allergen-coated Phazet lancets was higher than that of the cat allergen in-house reference extract (P = 0.048). There was a marked variation in potency between patients. Most of this variation was due to factors varying between patients, not between lancets, indicating that the composition of the allergen on the lancets was partly different from that of the standard.(ABSTRACT TRUNCATED AT 250 WORDS)

Standardization of mPEG-modified allergens.

Standardization of mPEG-modified allergens can be achieved by: standardization of allergens according to established principles; production of mPEG-modified allergens by a reproducible method; control of the batch-to-batch consistency by estimation of the allergenicity and the protein content in relation to the corresponding IHR and by determination of the degree of modification by estimation of the amount of PEG per unit allergen in the modified product. Standardization of mPEG-modified allergens should be looked upon only as a model for standardization of modified allergens. I hope that the authorities in Europe will set up strict and relevant criteria for the registration of similar products and also criteria for prolonging the registration of the older products without relevant standardization and documentation of safety and efficacy.

Precision of biological standardization of allergenic preparations.

Biological standardization (BS) aims at equilibration of the activity of allergen preparations from different types of allergen source materials. The biological unit (BU), proposed in the Nordic guidelines for 20 patients, has been found reproducible among different countries in Europe, but to be relatively imprecise, with a 95% confidence interval of about one power of 10. A more precise estimate of the biological activity of allergens or difference in sensitivity between populations would be of value. We used Ch, i.e. the concentration of allergen eliciting a wheal of the same size as histamine in the individual patient, estimated by regression line analysis. The Ch of 36 patients included in a BS trial was used. One of the 36 Ch-values was drawn randomly, and then sent back to the sample. This procedure was repeated 10, 20, 30, 40 and 60 times to create "samples" of different sizes. Ten samples of each size were produced. With 60 "individuals", the 95% confidence interval of the sample and the confidence interval of the medians were reduced to less than a factor of 2, i.e. to 74 to 128% of the median of the medians.

The skin prick test in the diagnosis of atopic allergy.

The factors that influence the results of skin prick tests are reviewed. The concentration and composition of allergen preparations, the test technique used, and the prevalence of allergy in the population studied are emphasized.

Mite allergens. Collection, determination, expression of results, and risk levels for sensitization and symptom induction.

The presence of mite allergens in dust can be determined by counting mites at different stages of development in dust and by determination of the major allergen content (Der p 1 ) in dust and air, which is crucially important to allergic patients. For comparison of results, similar methods for collection of dust and air must be used. Due to their size, mite bodies and fecal particles are airborne only directly after disturbance. Special filters should be used for dust collection, and upholstered surfaces should be vacuumed for 2 min/m2 (minimum 4 m2) and hard surfaces for 1 min/m2 (minimum 8 m2). Heavy contaminations should be removed. Preferably, the method given in the ISAAC study should be followed. Samples should be deep-frozen for at least some days to kill the mites. ELISA techniques, preferably using polyclonal antibodies and antigen with all isoforms present, should be used for determination of allergens. The allergen load has been given in ng/g of dust, but allergen/m2 or per sample area should be preferred. Allergen in the air should be given in pg/m3. A tentative limit of 2000 ng/g was proposed for sensitization and asthma. This limit is still valid on a population basis, but recent data indicate that highly susceptible young children become sensitized at concentrations 10-100 times lower and that ng levels of cat allergen/m3, as found in schools, induce chronic asthma.

Cross-reactivity between deciduous trees during immunotherapy. I. In vivo results.

Thirty-nine children with rhinoconjunctivitis due to birch pollinosis were given immunotherapy for 3 years with a potent, purified pollen preparation made from either birch alone or from a mixture of birch, alder and hazel. The therapy was evaluated with self-evaluation after each season, daily symptom score during the season, conjunctival provocation test and skin-prick test. All children but one considered that their condition had been improved by the treatment. Symptom scores decreased slightly more in the group treated with the mixture than in the group treated with birch only due to symptoms decreasing faster in the mixture treated group after each pollen peak (P less than 0.001). The sensitivity of the conjunctiva and the skin decreased significantly in both groups but without any significant differences between the groups. After 3 years the conjunctival sensitivity correlated strongly to eye symptoms during the season. Immunotherapy with a mixture of birch, alder and hazel appears to be at least as effective as a preparation made from birch only when treating birch pollinosis even in a region where birch is totally dominating among the pollens from deciduous trees.