屋尘螨与鸡蛋白交叉过敏的流行病学分析及实验研究

发布时间：2018-05-20 20:14

本文选题：交叉过敏 + 尘螨　；参考：《河北医科大学》2017年硕士论文

【摘要】：目的:过敏反应主要由血清中免疫球蛋白E(Ig E)介导,通过效应细胞释放生物活性介质而产生各种临床表现,如荨麻疹、血管神经性水肿等。有时,即使是微量蛋白质也可引起敏感个体出现致命的反应。随着对过敏性疾病的不断研究,交叉过敏反应也逐渐引起人们的重视。所谓交叉过敏就是当两种蛋白质在序列或结构上高度同源时,出现相同的抗原决定簇的几率增高,易出现交叉反应。检测屋尘螨与鸡蛋白之间是否存在交叉过敏、交叉过敏发生的概率以及发生交叉反应的蛋白种类;屋尘螨及鸡蛋白市售蛋白分子组分;比较免疫印迹法与狭缝免疫印迹法(Slot-blot)符合概率。基于目前屋尘螨与鸡蛋白过敏的发病率高,诊断及防治难的特点,本研究成果为屋尘螨与鸡蛋白过敏提供了可行且可靠的个体化治疗方法,并且使临床医师能更好的指导过敏患者正确饮食及生活习惯。方法:1流行病学调查及分析:对2015.01-2016.10就诊于河北医科大学第四医院皮肤科进行血清特异性过敏原检测的患者进行流行病学调查,并结合sIgE检测结果进行回顾性分析。2选取尘螨特异性sIgE(specific Ig E,sIgE)与总Ig E(total Ig E,t Ig E)同时阳性血清进行狭缝免疫印迹实验(Slot-blot),对比Slot-blot检测结果与艾康生物技术有限公司销售的免疫印迹法结果的一致性。3采用Slot-blot法进行交叉实验,采用鸡蛋白sIgE与t Ig E同时阳性血清与屋尘螨抗原进行反应,检测尘螨与鸡蛋白是否存在交叉反应及交叉概率。4采用Slot-blot进行抑制实验,先将鸡蛋白与屋尘螨sIgE阳性血清进行反应,然后再与包被的屋尘螨抗原反应,检测屋尘螨与鸡蛋白交叉抑制率和有效抑制浓度。5将屋尘螨及鸡蛋白粗提取液进行聚丙烯酰氨凝胶电泳(SDS-PAGE),检测各蛋白组分的完整性、分子量大小及所占比例。6对屋尘螨和鸡蛋白粗提取液进行免疫印迹实验(Western bolt,WB),选取屋尘螨sIgE与t Ig E同时阳性血清和鸡蛋白血sIgE与t Ig E同时阳性血清各10例为探针,检测各组分的抗原性。结果:1流行病学调查1.1本研究对2015.01-2016.10就诊于河北医科大学第四医院皮肤科的诊断为过敏反应相关性疾病的患者血清sIgE检测结果进行了回顾性分析。病例总数为5186例,男3100例,女2086例,年龄31.72±1.12岁,病程1周-数年,病种以荨麻疹、湿疹及皮炎为主。其中t Ig E阳性为3828例,尘螨sIgE阳性患者1223例,鸡蛋白sIgE阳性患者822例,其中尘螨与鸡蛋白sIgE同时阳性病例达419例,尘螨与鸡蛋白sIgE同时阳性病例占尘螨sIgE阳性的病例的34%。1.2过敏原sIgE阳性病例的分布情况:尘螨1223(31.9%),鸡蛋白822(21.5%),牛奶430(11.2%),牛羊肉344(9%),小麦223(5.8%),猫狗毛300(7.8%),霉菌172(4.5%),矮豚草166(4.3%),树组合114(3%),蟑螂109(2.8%),凬草96(2.5%),腰果花生159(4.2%),鱼虾蟹152(4%),芒果150(3.9%)。2免疫印迹法(艾康生物有限公司)与Slot-blot法实验结果对比分析60例尘螨sIgE阳性(免疫印迹法)的血清中,56例在Slot-blot实验中显示阳性,一致性概率达95%。3屋尘螨抗原与鸡蛋白阳性血清交叉过敏反应概率(Slot-blot法)将包埋于PVDF膜上的屋尘螨抗原与鸡蛋白sIgE阳性血清反应,随机检测60例,30例出现阳性反应,其出现交叉反应概率为50%。4鸡蛋白对屋尘螨抗原抗体反应的交叉抑制率的检测鸡蛋白对屋尘螨抗原抗体反应有明显的抑制作用,且呈浓度依赖性,当鸡蛋白干预抗原浓度为100μg/ml时,达到最大抑制率,为93%。5屋尘螨提取液与鸡蛋白提取液中各蛋白组分分析5.1屋尘螨列可见3条主要条带,分别是57kd,50kd,16kd,各组分所占比例依次是55.5%,43.4%,1.3%,其中57kd的蛋白所占比例最大。5.2鸡蛋白列可见4条主要条带,分别是75kd,44.5kd,28kd,14.3kd,各组分所占比例依次是19.6%,66%,7.7%,6.6%,其中所占比例最大的是44.5kd的条带。6屋尘螨与鸡蛋白免疫抗原性分析(WB检测)6.1 10例尘螨sIgE阳性患者的血清池为探针,50kd、57kd处可见明显阳性反应条带,考虑是Derp10(原肌凝蛋白)和Derp4(α-淀粉酶)。6.2 10例鸡蛋白sIgE阳性的患者血清池为探针。主要阳性反应条带有3条,分别是14.3kd、44.5kd、75kd,考虑分别为溶菌酶(Lys)、卵白蛋白(OVA)和卵转铁蛋白(OVT)。结论:1就诊于我院的过敏反应相关疾病的患者,主要过敏原是尘螨组合和鸡蛋白。2屋尘螨主要过敏原为Derp10(原肌凝蛋白)和Derp4(α-淀粉酶);鸡蛋白主要过敏原为溶菌酶(Lys)、卵白蛋白(OVA)和卵转铁蛋白(OVT)。3屋尘螨与鸡蛋白两种抗原之间存在相似的抗原表位,又存在各自特异性表位。4屋尘螨与鸡蛋白两种蛋白质由于存在相似抗原表位,而导致交叉过敏反应的发生。5鸡蛋白对屋尘螨的抗原抗体反应有交叉抑制作用,且抑制程度有浓度依赖性,当鸡蛋白浓度为100μg/ml时可达最大抑制,其抑制率为93%。
[Abstract]:Objective: Anaphylaxis is mainly mediated by immunoglobulin E (Ig E) in the serum, and produces various clinical manifestations, such as urticaria, vascular neuroedema, and so on. Sometimes, even trace proteins can cause a fatal response to sensitive individuals. Anaphylaxis is also attracting more and more attention. The so-called cross allergy is the increase in the probability of the same antigen determinant when two proteins are highly homologous in sequence or structure. Cross reaction is easy to occur. Protein species, house dust mite and chicken protein Market protein components; comparison between Western blot and slit immunoblotting (Slot-blot). Based on the high incidence of house dust mites and egg white hypersensitivity and difficult diagnosis and prevention, this study provides a feasible and reliable individualized treatment for house dust mites and egg white allergies. Methods: and enable clinicians to better guide the correct diet and living habits of allergic patients. Methods: 1 epidemiological investigation and analysis: 2015.01-2016.10 in the Department of Dermatology, the Department of Dermatology, the fourth hospital of Hebei Medical University, carried out the epidemiological investigation of the patients with specific allergen detection in the serum, and reviewed the results of the test in combination with the results of the test. Sex analysis.2 selected dust mites specific sIgE (specific Ig E, sIgE) and total Ig E (total Ig E, t) at the same time to carry out the slit immunoblotting experiment. The positive sera of white sIgE and t Ig E reacted with the house dust mite antigen to detect the cross reaction between the dust mite and the chicken protein and the cross probability of the chicken protein,.4 using Slot-blot to inhibit the test. First, the chicken protein was reacted with the positive serum of the house dust mite sIgE, and then it was reacted with the antigen of the house dust mites, and the cross inhibition of the house dust mites and chicken protein was detected. .5 and SDS-PAGE were carried out to extract the extract of house dust mite and egg white, and the integrality, molecular weight and proportion of.6 were detected by.6 in the immunoblotting test (Western bolt, WB) of house dust mite and egg white extract, and the positive serum of sIgE and t Ig E were selected as the positive serum. 10 cases of egg white blood sIgE and t Ig E positive serum were used as probes to detect the antigenicity of each component. Results: 1 epidemiological investigation was conducted in 1.1 studies on the retrospective analysis of the results of serum sIgE detection in patients diagnosed as allergic reaction related diseases in Department of Dermatology, Hebei Medical University, Hebei Medical University. The total number was 5186 cases, 3100 males and 2086 females, with age 31.72 + 1.12 years, and the course of disease was 1 weeks for 1 weeks. The disease species were urticaria, eczema and dermatitis. Among them, t Ig E positive was 3828 cases, dust mites sIgE positive patients were 1223, and egg white sIgE positive patients were 822 cases, including 419 cases of dust mite and chicken protein sIgE positive cases, dust mites and egg white sIgE simultaneously Yang The distribution of 34%.1.2 allergen positive cases in the cases of sIgE positive cases of dust mites were 1223 (31.9%), egg white 822 (21.5%), milk 430 (11.2%), beef and sheep meat 344 (9%), wheat 223 (5.8%), cat dog hair 300 (7.8%), fungus 172 (4.5%), dwarf ragweed, cockroach herb, cashew peanut 152 (4%), mango 150 (3.9%).2 immunoblotting (ikang bio limited company) and Slot-blot test results were compared and analyzed in the serum of 60 cases of sIgE positive (immunoblotting). 56 cases were positive in the Slot-blot test, and the consistency probability reached the probability of cross allergic reaction between 95%.3 house dust mite antigen and egg white positive serum (Slot-bl). Ot method) the reaction of house dust mite antigen embedded in PVDF membrane and positive serum of egg white sIgE was detected in 60 cases randomly and 30 cases were positive. The cross reaction probability of cross reaction was the inhibition rate of cross reaction of 50%.4 chicken protein to house dust mite antigen antibody reaction, and the chicken protein had obvious inhibition effect on the antibody reaction of house dust mite antigen. Degree dependence, when the concentration of chicken protein intervention antigen was 100 g/ml, the maximum inhibition rate was reached. 3 main bands were visible for the analysis of 5.1 house dust mite in the 93%.5 house dust mite extract and the chicken protein extract, respectively, 57kd, 50kd, 16kd, respectively. The proportion of each group was 55.5%, 43.4%, 1.3%, of which the 57kd protein accounted for the largest proportion. 5.2 egg white shows 4 main bands, which are 75kd, 44.5kd, 28kD, 14.3kd. The proportion of each group is 19.6%, 66%, 7.7%, 6.6%. The largest proportion is the 44.5kd strip.6 house dust mite and the chicken protein immune antigenicity analysis (WB test) 6.110 cases of dust mite sIgE positive patients' serum pool as the probe, 50kd, 57kd can be visible obvious The positive reaction strip was considered as the probe of the serum pool of Derp10 (primary myoprotein) and Derp4 (alpha amylase).6.2 10 cases of chicken protein sIgE positive. The main positive reaction strips were 3, respectively 14.3kd, 44.5kd, 75kd, respectively, which were considered to be lysozyme (Lys), egg white (OVA) and oval transferrin (OVT). Conclusion: 1 is the allergic reaction in our hospital. The main allergen of the related disease is the combination of dust mites and the main allergy of.2 house dust mites: Derp10 (procoagulin) and Derp4 (alpha amylase); the main allergy of egg white is lysozyme (Lys). There are similar epitopes between the two antigens of the ovalbumin (OVA) and the oval transferrin (OVT).3 house dust mite and egg white. The two proteins,.4 house dust mite and egg white in their specific epitopes, have similar antigen epitopes, which lead to cross anaphylaxis,.5 chicken protein has a cross inhibition effect on the antigen antibody response of house dust mite, and the inhibition degree is concentration dependent. When the chicken protein concentration is 100 g/ml, the maximum inhibition is reached and the inhibition rate is 93%..
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R593.1;R446.6

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