The Clinical Significance In Diagnosis Of Autoimmune Bullous Disease Through Combing Indirect Immnofluorescence On Salt-split Skin And Enzyme-linked Immunosorbent Assay

Objective: With the increasing aging degree of the population,the incidence of autoimmune bullous skin diseases,which is prone to middle-aged and elderly people,is gradually increasing.It seriously affects the quality of survival of patients.Early detection and accurate diagnosis are of great significance to the treatment and prognosis of the diseases.Autoimmune bullous skin diseases can be divided into two parts according the different pathsites,including the "intraepidermal blister disease"and"subepidermal blister disease.Because of the complex and diverse clinical manifestations,the existing detection methods,such as direct immunofluorescence(DIF)and enzyme-linked immunosorbent assay(ELISA),have some limitations in disease diagnosis and curative effect observation.Our study attempted to use the detection of indirect immno-fluorescence on salt-split skin(IIF-SSS)combined with the existing detection methods,through the judgment of the fluorescence deposition site and the expression of the specific circulating antibody Ig G subtypes in the blood,improved the sensitivity and specificity to the diagnosis of autoimmune bullous skin diseases,and facilitated the monitoring of disease progress.There are two advantages in our study,on the one hand,it can reduce the pressure and pain caused by repeated sampling of direct immunofluorescence;on the other hand,it can further improve the sensitivity and specificity of diagnosis by combining with enzyme-linked immunosorbent assay.Methods: Our study accumulated 25 patients of the intraepidermal blister disease(including of 18 cases of pemphigus vulgaris,4 cases of pemphigus foliaceus,2 cases of paraneoplastic pemphigus,1 case of pemphigus herpetiformis),26 patients of the subepidermal blister disease(including of 22 cases of bullous pemphigoid,1 case of linear Ig A bullous dermatosis,1 case of anti-P200 pemphigoid,1 case of herpes gestationis,1 case of epidermolysis bullosa acquisita).With the skin after plastic surgery as substrate,IIF-SSS method was used to detect the fluorescence deposition sites on the surface and dermis after splitting the substrate with 1mol/L Na Cl,and to classify autoimmune bullous skin disease.IIF-SSS was used to detect specific circulating antibody Ig G subtype in blood of patients and control group,and analyzed its expression degree in autoimmune bullous disease and its correlation with the severity of disease.Ig G subtype antibody was test by direct immuno-fluorescence(DIF)histopathology.ELISA was used to detect the expression of DSG1,DSG3,BP180 and BP230 antibodies in the blood of patients and control groups.Results:(1)IIF-SSS technique can effectively judge the type of special blister disease by the difference of fluorescence deposition position of epidermis and dermis.The IIF-SSS technique was used to detect specific antibody Ig G subtype,and the positive expression rates of Ig G1 and Ig G4 in serum of PV and BP patients were higher(Ig G1 positive rates were 55.56% and 55.56%,Ig G4 positive rate were 59.09% and 72.72% respectively).(2)According to Kappa test analysis,IIF-SSS and DIF had moderate and high consistency in the detection results of different antibody subtypes Ig G1,Ig G2,Ig G3 and Ig G4(KAPPA values were 0.556,0.469,0.577,0.675 respectively).(3)According to Spearman rank coefficient correlation analysis,the levels of Ig G1 and Ig G4 detected by IIF-SSS had good correlation with antibody titers detected by ELISA,and the correlation RS coefficients were 0.605 and 0.688 respectively.And Ig G1,Ig G4 were positively correlated with disease severity,and Ig G4 was more closely correlated.(4)Chi-square test was used to compare the sensitivity of DIF and IIF-SSS in diagnosing autoimmune bullous disease,which were 88.23%(95% CI: 79.1%-97.4%)and 88.23%(95% CI: 79.1%-97.4%)respectively.The specificity were 95%(95% CI:84.5%-105.5%)and 90%(95% CI: 75.65%-104.4%)respectively.(5)In different types of autoimmune bullous diseases,according to the detection results of serum antibodies by ELISA,the best critical points of sensitivity and specificity were found,and the diagnostic limit values were determined.The cases beyond the limit values were recorded as positive cases,and those below the limit values were recorded as negative cases.The number of cases was re-counted and accumulated in IIF-SSS grouping.The differences of sensitivity and specificity between IIF-SSS+ELISA and ELISA were statistically analyzed.The sensitivity of combined methods(89.47% of intraepidermal bullous disease,89.62% of subepidermal bullous disease)were higher than IIF(88.23%)and ELISA(88.23%).The specificity of combined methods(92.5%for both intraepidermal bullous disease and raepidermal bullous disease)were slightly lower than IIF(95%),meanwhile slightly higher than ELISA(90%).Conclusions:(1)IIF-SSS can identify intraepidermal blister disease and subepidermal blister disease by the specificity of antibody deposition site.(2)The detection of circulating specific antibodies by IIF-SSS provided an important basis for the evaluation of autoimmune bullous disease.Specific antibodies Ig G1 and Ig G4 were positively correlated with the severity of autoimmune bullous disease,and Ig G4 was more closely correlated.(3)The evaluation of IIF-SSS and DIF on the detection results of different antibody subtypes Ig G1,Ig G2,Ig G3 and Ig G4 has medium and high consistency.Compared to DIF,IIF-SSS was convenient and easily acceptable to patients.(4)Compared with ELISA,IIF-SSS combined with ELISA has higher sensitivity and specificity,and the position of circulating antibody fluorescence deposition was more specific for diagnosis.