Immunoblot Analysis of Hypoallergenic Buckwheat with Monoclonal Antibodies to Raw Buckwheat / 소아알레르기및호흡기학회지

BACKGROUND AND OBJECTIVE: It is generally accepted that one third of children with food allergy lose their clinical activity if the responsible food allergen can be identified and completely eliminated from their diet. But buckwheat allergy rarely lose clinical activity. So, we have produced hypoallergenic buckwheat(HBW) according to Ikezawa's method, and characterized the allergenicity by immunoblot method using monoclonal antibodies to raw buckwheat. MATERIAL AND METHODS: HBW was prepared from raw buckwheat by salting with 0.5 M NaCl. 18 monoclonal antibodies to raw buckwheat were made according to standard polyehtylene glycol method. We performed SDS-PAGE of HBW and transferrd to nitrocellulose membranes, which were immunoblotted by selected 3 monoclonal antibodies to raw buckwheat. RESULTS: The protein concentration of HBW was 47.3 microgram/microliter. As the result of SDS-PAGE of HBW the protein bands of 50, 36, 13 kD of raw buckwheat disappeared. 3 monoclonal antibodies were selected due to their higher specificities in ELISA. Immunoblot study showed that 2 among 3 monoclonal antibodies bind to HBW. CONCLUSION: The SDS-PAGE of HBW showed less protein bands in comparison to raw buckwheat. But the allergic component of HBW still remained. So further studies are required to eliminate the residual allergenicity of hypoallergenic food.

A Clinical Study on Oral Milk Provocation and Elimination Test / 소아알레르기및호흡기학회지

PURPOSE: A Cow's milk allergy (CMA) is one of common problems during first year of life. Though milk caused allergic problems, their parents didn't know it, and gave milk to their children. But the parents don't give milk to their children if they know it. Therefore it may cause problems of growth and development in children. The purpose of this study is to analyze those clinical features of milk allergy confirmed by oral provocation test. METHODS: We carried out the retrospective study on 21 patients who had been performed oral milk provocation test at Yonsei university children's allergic clinic. RESULTS: 9 out of 21cases (43%) showed positive milk oral provocation test. The most common clinical symptom & sign was urticaria, followed by respiratory symptoms and abdominal symptoms. The rate which past history and positive skin test corresponded to oral provocation test was very high (100%). CONCLUSION: The most valuable diagnosis of food allergy is oral provocation test. However, predictive value of allergy skin test and past history was very high in milk allergy.

A Study on Provocation Test by Cat Contacts / 소아알레르기및호흡기학회지

PURPOSE: Cat is the pet which many of us quite frequently contact without any specific precautions. According to the allergy skin tests conducted in children, cat antigen shows a comparatively high positive reaction, and past history also shows that there are many patients who suffer allergies from cat contacts. Therefore, once it is diagnosed as an allergy caused by cat, it is important that patients be isolated in order to restrict their contact with cat. In this context, this study was conducted for the purpose of finding out how serious and how frequent the allergies by cat contacts are. METHODS: Forty five children patients who visited to the Allergy Clinic for Children, Severance Hospital annexed to the Medical College, Yonsei University were enrolled in this study. Some of patients had shown a positive reaction against cat antigen in the allergy skin test, and others had a past history of developing symtoms when contacted by cat. The patients were isolated from cat for seven days, and then contacted with cats for 30 minutes and observed the provocated symptoms. RESULTS: 1) Among the tested patients, the number of cases who were provocated by the symptoms after cat contact were 18 (40%). 2) Among those patients who were shown to be positive only in allergy skin test, only 24.2% proved to be provocated by the symptoms, while 90.0% of those who had a past history in addition to allergy skin test proved to be provocated by the symptoms. 3) Symptoms provocated after cat contact showed allergic rhinitis patients 88.2% which was the highest ratio, bronchical asthma, 35.3%, urticaria, 29.4%, and allergic conjunctivitis, 23.5%, respectively. 4) The onset and duration of symptoms provcated were under 10 minutes and from 30 to 60 minutes, which included most cases in the test. 5) Total IgE and eosinophil count of patients did not show any meaningful differences between the group which was provocated and that which was not provocated. CONCLUSION: It was shown that 40% patients who were suspected of cat allergy had a positive reaction to the cat contact provocation test. Ninety percents of patients who showed both positive allergy skin test and past history were provocated after cat contact.

Identification and Characterization of German Cockroach Allergen / 소아알레르기및호흡기학회지

PURPOSE: Cockroaches have been reported one of the major causes of allergic disease such as bronchial asthma and allergic rhinitis. About 10% of children and 20% of adult respiratory allergy patients have positive results of allergy skin tests to cockroach allergen. This finding suggests that cockroach is also important allergen in Korea. Among 4,000 species, German cockroach seems predominant species in the Korean peninsula. Identification and charactrerization of the major allergen is the first step in German cockroach allergen research. But, there have been controversies. It may depends upon different identifying methods. The aim of our study was to identify the major allergen in German cockroach using Westernblot. METHODS: Crude German cockroach allergen extracts with or without protease inhibitor cocktail mixture were prepared, and protein patterns of these two extracts were compared using SDS-PAGE and Coommssie stain. Sera from 32 atopic asthmatic patients with positive allergy skin test against cockroach allergen were used for IgE immunoblot. RESULTS: Coomassie stain after SDS-PAGE demonstrated some discordance between the differently prepared crude allergen extracts. In comparison with the allergen extract which abolished endogenous protease activity, the untreated extract revealed new 32, 55 and 78 kD protein bands and 41 kD protein band was disappeared. About 60% of sera with positive allergy skin test revealed IgE binding bands in Westernblot. Among them, 10, 21, 25, 36 and 52 kD protein band might seemed important German cockroach allergen. CONCLUSION: Finding above may suggest that endogenous protease could denature allergenic proteins in the process of crude cockroach allergen extract. The 10, 21, 25, 36, and 52 kD might be the important allergens in German cockroach.

Production of Recombinant Buckwheat Allergen / 소아알레르기및호흡기학회지

PURPOSE: Buckwheat is one of the most common food allegen in Korea and frequently elicit severe allergic reactions. However, up to now, only few reports on buckwheat allergens have been reproted. The purpose of this study was to isolate mRNA for the production of recombinant buckwheat allergens. METHODS: After the isolation of mRNA from ripening buckwheat seeds, in vitro translation was performed. The proteim patters of in vitro traslate products were identified using SDS-PAGE. RESULTS: A260/A260 ratio total RNA was 1.9 and mRNA was 2.0. In vitro translate products showed towered molecular buckwheat proteins such as 1, 3, and 14kD, while other high molecular weight protein of buckwheat seed were not shown in SDS-PAGE. CONCLUSION: mRNA of buckwheat was purified only from ripening seeds. The composition of mRNA was different according to the ripening periods. It is believed that this finding can give a clue to the basic research of buckwheat allergen.

A Case of Kawasaki Disease of Five Recurrences During 4 Years

Kawasaki disease is an acute illness of early childhood, especially less than 5 years of age, characterized by prolonged fever (>5 days), changes of lips and oral cavity, changes of peripheral extremities, polymorphous skin rash, bilateral conjunctival congestion and acute nonpurulent cervical lymphadenopathy. Usually, the recurrence of Kawasaki disease is not frequent. It is thought that the factors in recurrence of Kawasaki disease was sex, age and IV gammaglobulin. We have experienced the first attack of Kawasaki disease at 18 months-old female who had 5 recurrences during 4 years.

Isolation of Enterovirus in Patients with Aseptic Meningitis in Spring, 1996 / 감염

BACKGROUND: Aseptic meningitis is an acute viral infection of the central nervous system that occurs commonly in childhood. Although the etiologic agent is not always identified, the human enteroviruses are responsible for most cases of aseptic meningitis in which a cause can be identified. Enterovirus causes approximately 80% of all cases of aseptic meningitis. In 1993, there was a nationwide epidemic of aseptic meningitis by echovirus 9 and 30. We reported that the cause of aseptic meningitis in 1994 was echovirus 3 and coxsackievirus B3 and echovirus 7 in 1995. This study was done to detect the causative agent of aseptic meningitis in spring, 1996. METHODS: To isolate the causative viruses, stool and cerebrospinal fluid specimens from the patients with aseptic meningitis, who were admitted to Severance Hospital in 1996, were collected. Cultured RD cells and HEp-2 cells were inoculated with specimens to see the cytopathic effects. Neutralizing antibody tests using enterovirus serum pool were done on the specimens with the cytopathic effects. RNA was isolated from the cultured supernatants of the infected cells. Oligonucleotide was synthesized by PCR, which was run on polyacrylamide gel after purification with HPLC. After running the DNA produced by using Geneamp RNA PCR kit, electrophoresis was done. RESULTS: Enteroviruses were isolated from 14 out of 17 patients. Among these fourteen, Coxsackievirus B1 was isolated in 13 patients and poliovirus in one patient. PCR product from these viruses showed a 152bp band on electrophoresis. CONCLUSION: The causative virus of aseptic meningitis in patients who were admitted to Severance Hospital during the spring season of 1996 was Coxsackievirus B1.