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Figure 1 VGCC antibody uptake in cerebellar slice culture

Published byΆργος Παπακώστας Modified over 5 years ago

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1 Figure 1 VGCC antibody uptake in cerebellar slice culture
VGCC antibody uptake in cerebellar slice culture Slice culture of rat cerebellum incubated with patient serum containing VGCC autoantibodies and examined at different points in time. (A) Slice culture incubated with normal serum for 144 hours, treated with SYTOX dyes to detect cell death, and labeled with Cy5-conjugated anti-human IgG, showing absence both of neuronal antibody binding and of SYTOX staining indicative of cell death. (B, C) Slice cultures incubated for 48 and 72 hours with serum containing VGCC autoantibodies. In these images, binding of the patient's IgG is labeled red; rare dead cells outside the Purkinje cell layer are labeled green. At 48 hours, IgG can be detected throughout the neuropil with labeling of occasional Purkinje cells (arrows) (B). By 72 hours, there is extensive binding of IgG to Purkinje cell membranes (arrows) (C). (D–F) A slice culture incubated with patient serum for 144 hours and labeled with Cy5-conjugated anti-human IgG (D), stained with SYTOX dyes to detect cell death (E), and as a merged image (F). In cultures at this time point, some Purkinje cells continued to show surface binding of antibody panels (G–I). In other areas, however, Purkinje cells showed uptake of IgG with cytoplasmic and nuclear labeling (D); some IgG-containing Purkinje cells demonstrated staining with SYTOX dyes, indicating cell membrane disruption and death (E, F), whereas others excluded SYTOX dyes, indicating antibody uptake by viable Purkinje cells. Purkinje cell antibody binding and cell death at 144 hours are shown at higher power in panels G through I. Arrows indicate the same cells in panels D–F and in G–I. Scale bars = 20 μm. IgG = immunoglobulin G; VGCC = voltage-gated calcium channel. Marilyn McKasson et al. Neurol Neuroimmunol Neuroinflamm 2016;3:e222 © 2016 American Academy of Neurology

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