rat brain sections  (Vector Laboratories)

Journal: Frontiers in Immunology

Article Title: Antibody induced seizure susceptibility and impaired cognitive performance in a passive transfer rat model of autoimmune encephalitis

doi: 10.3389/fimmu.2023.1268986

Figure Lengend Snippet: Experimental design. Experimental protocol to assess the role of NMDAR and LGI1 antibodies in in a passive transfer rat model of autoimmune encephalitis (AE). Model based on the autoantibodies obtained from blood samples of AE patients (A) . Total IgG obtained from the pooled sera of NMDAR and LGI1 antibody positive patients with epileptic seizures and healthy subjects was purified (B) and applied chronically every other day for 11 days into the cerebral lateral ventricle. Antibody infusions were performed on day 11, 13, 15 and 17 and 19 (as presented green labelled ICV). On days without infusion (10, 12, 14, 16, 18, 20th days, as presented blue). (C) . Spontaneous seizure development was followed by recording cortical electroencephalography (EEG) on consecutive days and after infusion administration for 2h. Behavioral tests for memory (novel object recognition test (NOR) and Y-maze) and locomotor activity (open field and rotarod tests) were applied to the animals before the antibody infusions started and at the end of the infusions. Then, pentylenetetrazole (PTZ) was administered at a convulsive dose (45 mg/kg) intraperitoneally in order to detect possible changes in seizure susceptibility. Immunohistochemistry was done for assessment of hippocampal astrocyte proliferation and expression intensity of target NMDAR and LGI1 antigens. Coronal brain section showing the appropriate ventricular placement of the guide cannullae (D) . Created with Biorender.com .

Article Snippet: To ensure the diffusion of human antibodies in the hippocampus, frozen rat brain sections were incubated with biotinylated anti-human-IgG antibodies (Vector Laboratories, Newark, CA, USA), followed by the avidin-peroxidase and diaminobenzidine ( – ) ( ).

Techniques: Purification, Activity Assay, Immunohistochemistry, Expressing

Journal: Frontiers in Immunology

Article Title: Antibody induced seizure susceptibility and impaired cognitive performance in a passive transfer rat model of autoimmune encephalitis

doi: 10.3389/fimmu.2023.1268986

Figure Lengend Snippet: Antibody treatment induced astrocyte proliferation and NMDAR loss in the hippocampus Immunohistochemical staining of frozen sections (A) of rats treated icv with NMDAR-IgG (left column), LGI1-IgG (middle column) containing total serum IgG and healthy control (HC)-IgG (right column) using commercial antibodies directed against glial fibrillary acidic protein (anti-GFAP, upper row, brown color), anti-NMDAR (middle row, brown color) and anti-LGI1 (lower row, green color). Counter-staining was done with hematoxylin (blue color) in GFAP and NMDAR assays and DAPI (blue color) in LGI1 assays. Original magnification is 20x for upper row and 4x for middle and lower rows. Quantification of immunohistochemical stainings by Image J analysis (B) : GFAP, NMDAR and LGI1. Data were expressed as mean ± SEM p value denoted on the upper left corner of the panel is obtained by ANOVA; ***p<0.001 by Tukey’s post-hoc test.

Article Snippet: To ensure the diffusion of human antibodies in the hippocampus, frozen rat brain sections were incubated with biotinylated anti-human-IgG antibodies (Vector Laboratories, Newark, CA, USA), followed by the avidin-peroxidase and diaminobenzidine ( – ) ( ).

Techniques: Immunohistochemical staining, Staining, Control