PMC9262380_01
Antibody diagnostics. Immunofluorescence studies: diluted patient material is incubated with mouse brain sections or human embryonic kidney (HEK) cells transfected with the antigens of interest. Binding of patient antibodies to the respective matrix is visualized by a secondary anti-human/canine antibody coupled with an immunofluorescence dye resulting in a red signal. (A-C) Tissue-based assays, mouse cerebellum. (D) cell-based assay with HEK cells which were transfected with the gamma-aminobutyric acid (GABA)-A receptor (GABAAR) subunits alpha1beta3 (all assays from Euroimmun, Lubeck, Germany). Molecular layer (ML), purkinje cell layer (Pu), granular cell layer (GL), white matter (WM). (A) Human antibodies against the GABAAR. (B) Human antibodies against the GABABR. (C) The canine antibodies against the GABAAR. Please note that the binding pattern is similar to (A) and not to (B). (D) The canine serum binds to GABAAR transfected HEK cells. Nuclear counterstaining with Hoechst 33342 in blue. Bar in (A), valid also for (B,C): 100 mum. Bar in (D): 25 mum.
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F1
An extensive antibody profile demonstrated a neuropil staining pattern on a tissue-based assay compatible with GABAAR antibodies (Figure 1); the diagnosis was confirmed by binding of serum and CSF to GABAAR (alpha1beta3 subunits) transfected Human Embryonic Kidney cells [all assays from Euroimmun, Lubeck, Germany, used with in-house-protocols; for the canine material, secondary anti-dog Immunoglobulin G (IgG) was used].