Sensitive and specific assays for routine serological diagnosis of epidermolysis bullosa acquisita
Authors
Lars Komorowski ; Ralf Müller ; Artem Vorobyev ; Christian Probst ; Andreas Recke ; Marcel F. Jonkman ; Takashi Hashimoto ; Soo-Chan Kim ; Richard Groves ; Ralf J. Ludwig ; Detlef Zillikens ; Winfried Stöcker ; Enno Schmidt
Citation
JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY, Vol.68(3) : 89-95, 2013
autoantibody ; ELISA ; immunofluorescence ; type VII collagen
Abstract
BACKGROUND:
Epidermolysis bullosa acquisita (EBA) is a severe autoimmune subepidermal blistering disease characterized by autoantibodies against the N-terminal collagenous domain (NC1) of type VII collagen (Col VII).
OBJECTIVE:
Development of reliable assays for the detection of anti-Col VII-NC1 antibodies.
METHODS:
NC1 was expressed in human HEK293 cells and used as target antigen in an enzyme-linked immunosorbent assay (ELISA) and in an immunofluorescence assay (IFA). These two assays were probed in a large cohort of patients with EBA (n = 73), bullous pemphigoid (BP, n = 72), anti-p200 pemphigoid (n = 24), anti-laminin 332 mucous membrane pemphigoid (MMP, n = 15), pemphigus vulgaris (PV, n = 24), and healthy control subjects (n = 254).
RESULTS:
The cut-off for the ELISA was optimized for accuracy by receiver-operating characteristics (area under the curve [AUC] = 0.9952). IgG reactivity against NC1 was detected in 69 of 73 EBA (94.5%) and 5 control sera (2 healthy controls and 3 BP patients), resulting in a specificity of 98.7%. The IFA showed a sensitivity of 91.8% and specificity of 99.8%. Reproducibility of the ELISA was demonstrated by an intra-class correlation coefficient of 0.97. IgG subclass analyses by ELISA revealed IgG1, IgG2, IgG3, and IgG4 anti-NC1 reactivity in 83.6%, 85.3%, 37.7%, and 83.6% of EBA sera, respectively.
LIMITATIONS:
The novel assays were not evaluated prospectively and their use in monitoring serum levels during the disease course was not tested.
CONCLUSION:
The two assays are highly specific and sensitive to diagnose EBA. Their diagnostic competence was demonstrated in a large cohort of well-characterized EBA sera.