Low sensitivity of type VII collagen enzyme-linked immunosorbent assay in epidermolysis bullosa acquisita: serration pattern analysis on skin biopsy is required for diagnosis.

The type VII collagen (coll VII) enzyme-linked immunosorbent assay (ELISA) has been reported to have high sensitivity (> 93%) and specificity (> 96%) for diagnosing epidermolysis bullosa acquisita (EBA) in patients who are seropositive on indirect immunofluorescence on salt-split skin (SSS). To investigate the added value of the coll VII ELISA in the laboratory diagnosis of SSS-positive and SSS-negative EBA and to correlate the ELISA index with disease episode. The coll VII ELISA was performed on banked sera of 28 patients with EBA: 15 SSS positive and 13 SSS negative. Sera from healthy blood donors (n = 17) and patients with other autoimmune blistering diseases (n = 29) served as controls. In four patients, the ELISA index was measured longitudinally. Serration pattern analysis by direct immunofluorescence has been prospectively performed since 2000 in 19 patients. The sensitivity in the SSS-positive group was 80% whereas it was 23% in the SSS-negative group. In the prospective EBA subset it was 45%. The sensitivity of u-serration pattern analysis on skin biopsy was 89%. Ten (53%) of these cases were seronegative with both ELISA and SSS, and would have been missed by serum analysis alone. Of the 46 control sera, one serum tested positive (specificity 97·8%). The coll VII ELISA correlated with disease activity over time in individual patients. The coll VII ELISA has limited added value in SSS-negative EBA cases. The ELISA test is valuable in differentiating EBA from antilaminin-332 mucous membrane pemphigoid and anti-p200 pemphigoid and in its ability to monitor patients with EBA serologically. U-serration pattern analysis on immunofluorescence skin biopsy is the gold standard for the diagnosis of EBA.

Terra JB ，Jonkman MF ，Diercks GF ，Pas HH ... -  《-》

Diagnosis and disease severity assessment of epidermolysis bullosa acquisita by ELISA for anti-type VII collagen autoantibodies: an Italian multicentre study.

Epidermolysis bullosa acquisita (EBA) is a rare autoimmune mucocutaneous bullous disease caused by autoantibodies against type VII collagen, a component of anchoring fibrils that stabilizes dermoepidermal adherence. Type VII collagen is composed of a collagenous domain linked by the noncollagenous (NC)1 and NC2 domains.  To assess the repeatability, sensitivity and specificity of a recently developed enzyme-linked immunosorbent assay (ELISA) for detection of anti-type VII collagen autoantibodies, and to ascertain whether they may be a marker of disease activity in EBA. Using this ELISA, which was able to recognize autoantibodies against the NC1 and NC2 epitopes of type VII collagen, we tested 14 EBA sera, 30 healthy control sera and 113 disease control sera. In the EBA sera group, 12 out of the 14 samples were positive in ELISA, with autoantibody titres varying from 7·2 to 127·9UmL(-1) (cutoff value <6), the sensitivity of the method being 86%. Among the controls, only two bullous pemphigoid sera tested positive, the specificity being 98·6%. A good correlation was found between EBA disease severity, expressed as autoimmune bullous skin disorder intensity score, and the serum levels of anti-collagen VII autoantibodies, measured by ELISA (n =14; r=0·965; P=0·0001). The intra- and interassay coefficients of variation of the ELISA method ranged from 6·3% to 18·3%.  This NC1+NC2 ELISA can be a practical assay for the diagnosis of EBA. The correlation between autoantibody titres and disease severity suggests its usefulness as a marker of disease activity in EBA However, this should be confirmed by studies on larger series of patients.

Marzano AV ，Cozzani E ，Fanoni D ，De Pità O ，Vassallo C ，Berti E ，Parodi A ，Crosti C ，Cugno M ... -  《-》

Serum levels of anti-type VII collagen antibodies detected by enzyme-linked immunosorbent assay in patients with epidermolysis bullosa acquisita are correlated with the severity of skin lesions.

Epidermolysis bullosa acquisita (EBA) is a chronic autoimmune subepidermal bullous disease characterized by circulating autoantibodies against type VII collagen. Detecting these autoantibodies is crucial for the diagnosis of this disease, and is also useful for measuring disease activity. Enzyme-linked immunosorbent assay (ELISA), a quantitative method to measure anti-type VII collagen antibody levels, is currently available to diagnose EBA. The aim of this study was to investigate the relationship of ELISA with overall clinical severity. Sera from patients with EBA (n = 30), bullous pemphigoid (n = 20), anti-laminin γ1 pemphigoid (n = 9) and healthy donors (n = 24) were tested using ELISA, using the recombinant non-collagenous 1 (NC1) and 2 (NC2) domains of type VII collagen. Relationships between clinical characteristics, indirect immunofluoroscence (IIF) titres and ELISA values were investigated. The sensitivity and specificity of the EBA ELISA were 96.7% and 98.1%, respectively. There was no significant difference between ELISA results for classic and inflammatory types. The severity of skin involvement was positively correlated with both ELISA value (r = 0.87, P < 0.01) and IIF titre (r = 0.59, P < 0.01). Time sequence analysis in four patients with EBA showed that ELISA values reflect disease activity better than IIF titres. Type VII collagen ELISA using the NC1 and NC2 domains is useful for diagnosing EBA and monitoring disease severity.

Kim JH ，Kim YH ，Kim S ，Noh EB ，Kim SE ，Vorobyev A ，Schmidt E ，Zillikens D ，Kim SC ... -  《-》

Comparison of 3 type VII collagen (C7) assays for serologic diagnosis of epidermolysis bullosa acquisita (EBA).

Serologic diagnosis of epidermolysis bullosa acquisita (EBA) relies on the detection of circulating autoantibodies to type VII collagen (C7). We sought to compare the diagnostic performances of a commercialized enzyme-linked immunosorbent assay (ELISA) using C7 noncollagenous (NC) domains (C7-NC1/NC2 ELISA) and indirect immunofluorescence (IIF) biochip test on NC1-C7-expressing transfected cells (IIFT), with a full-length-C7 ELISA developed in our laboratory. C7-NC1/NC2 ELISA, IIFT, and full-length-C7 ELISA were run on 77 nonselected consecutive EBA sera. C7-NC1/NC2 ELISA, IIFT, and full-length-C7 ELISA were positive, respectively, for: 30%, 27%, and 65% of the 77 sera; 43%, 32%, and 80% of 44 sera labeling the salt-split-skin (SSS) floor (F) by IIF (SSS/F(+)); 9%, 22%, and 47% of 32 SSS/F(-) sera; 28%, 28%, and 58% of classic EBA; 41%, 41%, and 82% of inflammatory EBA; and 18%, 0%, and 55% of mucous-membrane-predominant EBA. Significant differences for all sera were found between: the 2 ELISAs for the 77 sera, SSS/F(+) and SSS/F(-) sera, and IIFT versus full-length-C7 ELISA. The retrospective design was a limitation. C7-NC1/NC2 ELISA and IIFT sensitivities for serologic diagnoses of EBA were low. Full-length-C7 ELISA was significantly more sensitive and could serve as a reference test.

Seta V ，Aucouturier F ，Bonnefoy J ，Le Roux-Villet C ，Pendaries V ，Alexandre M ，Grootenboer-Mignot S ，Heller M ，Lièvre N ，Laroche L ，Caux F ，Titeux M ，Hovnanian A ，Prost-Squarcioni C ... -  《-》

Anti-type VII collagen autoantibodies, detected by enzyme-linked immunosorbent assay, fluctuate in parallel with clinical severity in patients with epidermolysis bullosa acquisita.

Epidermolysis bullosa acquisita (EBA) is an autoimmune subepidermal blistering disease caused by autoantibodies against type VII collagen. An enzyme-linked immunosorbent assay (ELISA) is currently available to detect autoantibodies in EBA. There have been reports suggesting generically that ELISA indices reflect EBA disease severity; however, there is, as yet, no conclusion as to whether ELISA indices fluctuate with disease activity over time in each EBA patient. This study aimed to investigate whether ELISA titers fluctuate with EBA disease activity and to validate the clinical significance of checking ELISA values in EBA by monitoring type VII collagen ELISA titers and disease severity, evaluated in terms of numbers of blisters and erosions as a clinical score, over time in three Japanese patients with EBA. All three cases in this study, which were treated successfully, showed titers of anti-type VII collagen autoantibodies detected by ELISA that fluctuated in parallel with disease activity. Especially in case 1, we could determine that the expanding erosions were not due to flare-ups of EBA because the ELISA indices stayed low, although new lesions continued to appear. In fact, control of infection and nutrition helped the lesions to become epithelialized. In conclusion, we found that repeated ELISA measurements are useful in monitoring disease activity and making decisions in EBA treatment plans.

Ito Y ，Kasai H ，Yoshida T ，Saleh MA ，Amagai M ，Yamagami J ... -  《-》