| Objective: In recent decades,the incidence of allergic diseases is increasing year by year,which not only affects the quality of work and life,but also brings huge economic burden to individuals,families and society.In order to correctly diagnose and control allergic diseases,a variety of allergen detection methods have emerged.Among them,the detection of serum specific IgE(sIgE),as a safe,effective and convenient detection method,is widely used in the diagnosis of allergic diseases.However,the inconsistence of the clinical and laboratory diagnosis is founded sometimes.In order to optimize the serum house dust mite sIgE detection system and reduce the occurrence of false-negative and false-positive results,we foucus on the Dermatophagoides pteronyssinus and designe the following schemes using molecular biology methods.Methods:1 Epidemiological investigation: an epidemiological investigation was performed on patients with allergic-related diseases in the Department of Dermatology,the Fourth Hospital of Hebei Medical University from January2017 to December 2017,who were carried out the detection of s IgE.2 Serums of patients,who had the inconsistent result of serum s IgE test and clinical manifestations for house dust mite,were selected for the slot-blot(SB)test of extract of Dermatophagoides pteronyssinus.We compared the results of slot-blot test and serum s IgE detection system.3 Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE)and western-blot(WB)test were used to analyze the components of Dermatophagoides pteronyssinus and their allergic reaction.4 Further analysis of the Dermatophagoides pteronyssinus were performanced by high performance liquid chromatography(HPLC),followedby SDS-PAGE and WB test.5 The results of the main peak 1 by SB test was compared to that of serum sIgE detection system.Results:1.The main allergens of patients visiting in our hospital were house dust mite,egg white,milk,beef and mutton.2.Epidemiological survey results: Totally there were 2,070 patients registered,including 817 males and 1253 females,and the mean age is29.8±7.8.By comparing the results of epidemiological investigations and the serum sIgE detection system,we found that 182 patients with suspected dust mite allergy were inconsistent with serum sIgE test results.3.The accuracy of SB test for extract of Dermatophagoides pteronyssinus is 35.2% higher than that of serum sIgE detection system.4.By SDS-PAGE and WB,so many bands were founded in the extract of Dermatophagoides pteronyssinus.The bands of 48.7kDa,33.3kDa-29.8kDa,and 17.6kDa were more obvious,considering as Der p 10,Der p1,Der p 3,Der p 9,and Der p 2,respectively.5.Seven main peaks were detected by HPLC chromatogram for the extract of Dermatophagoides pteronyssinus.By analyzing the results of SDS-PAGE and WB test,it can be seen that the more obvious reaction bands were 210.9kDa,129.2kDa-102.3kDa,33.6kDa,and 27.8kDa,considering as Der p 14,Der p 11,Der p 15,Der p 9,Der p 3,Der p 6,Der p 7,and Der p 8.6.The main peak 1 SB test can improve the coincidence rate of the serum allergen sIgE detection(49.4%).7.HPLC technology and SB test can be used to optimize the detection method of dust mite allergen.Conclusions:1.The main allergens are house dust mite,egg whites,milk,beef and mutton in the patients visiting in our hospital.2.The result by SB test for the extract of Dermatophagoides pteronyssinus can improve the accuracy of the current serum sIgE detection(35.2%).3.The major allergens of Dermatophagoides pteronyssinus were considered to be Der p 1,Der p 2,Der p 3,Der p 9 and Der p 10.4.The major allergens of main peak 1 may be Der p 3,Der p 6,Der p 7,Der p 8,Der p 9,Der p 11,Der p 15,and Der p 14.5.The main peak 1 SB can improve the false negative result of the current sIgE detection method,but its guiding significance for diagnosis needs further study.6.HPLC technology and SB test can be used to optimize the detection method of dust mite allergen. |
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