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Khwaja Aftab Rashid, C. Stephen Foster, A. Razzaque Ahmed; Identification of Epitopes Within Integrin β4 for Binding of Auto-Antibodies in Ocular Cicatricial and Mucous Membrane Pemphigoid: Preliminary Report. Invest. Ophthalmol. Vis. Sci. 2013;54(12):7707-7716. doi: https://doi.org/10.1167/iovs.12-11404.
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© ARVO (1962-2015); The Authors (2016-present)
To identify the epitopes on human β4 integrin to which the sera of patients with ocular cicatricial pemphigoid (OCP) and mucous membrane pemphigoid (MMP) without ocular involvement bind.
Fragments of the intracellular domain of the β4 molecule were cloned, expressed, purified and peptides were synthesized. Antibodies to various fragments and peptides were produced in rabbits. Binding specificity was determined via Western blot and blocking experiments. Test sera and controls were injected into neonatal BALB/c mice for in vivo passive transfer.
Sera from patients with OCP, MMP, and both OCP and MMP were bound to cloned fragments of IC3.0. Its subcloned fragments IC3.4 (1489 aa–1572 aa) and IC3.4.1 (1489 aa–1510 aa) were bound with the sera from patients with OCP only. Subcloned fragments IC3.6 (1573 aa–1822 aa) and IC3.6.1 (1689 aa–1702 aa) were bound with MMP sera only. No cross-reactivity in binding was observed. Immuno-affinity–purified sera from patients with OCP, MMP, and rabbit antibodies to IC3.0, IC3.4, IC3.4.1, IC3.6, and IC3.6.1, when injected in neonatal BALB/c mice, produced subepidermal blisters in their skin.
These preliminary observations identified IC3.4.1 as the possible epitope for the binding of OCP auto-antibody and IC3.6.1 as the possible epitope for the binding of MMP auto-antibody without ocular disease. Antibodies specific to these peptides produced blisters when injected in mice. Still-unidentified epitopes may exist. These observations may enhance our understanding of the role of β4 integrin in the pathobiology of OCP and MMP. Early diagnosis may be possible if serologic tests with specificity and sensitivity can be developed.
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