Fig. 1

Methodical procedure of morphometric analysis of p-α-syn positive aggregates in neuronal somata of submucosal ganglia. a Screening of submucosal ganglia in orthogonal sections of a deep submucosal biopsy containing the mucosal and submucosal layer. Ganglia (arrowhead in (a) and inlet of a) and nerve fibers of the submucosal plexus were identified by conventional immunohistochemistry with the pan-neuronal marker PGP 9.5. b Dual-label immunohistochemistry with PGP 9.5 (red) and p-α-syn (green) of a submucosal ganglion. Number and area of neurons detected by the pan-neuronal marker PGP 9.5 were recorded (c, blue circles). d Insert from c. Total number and area of p-α-syn positive structures per neuron was calculated in photographs displayed in reverse grey-scale mode. p-α-syn positive aggregates were subdivided into three groups according to size (L = large size, M = medium size, S = small size). Nuclear counterstain with DAPI (blue), bars (a): 500 μm, inlet in (a) = 100 μm: (b-d): 20 μm. e To determine the threshold between medium and large size p-α-syn positive aggregates the crossing point of frequency distribution curves of p-α-syn positive aggregates in patients with PD (blue squares) and controls (red triangles) was defined (black arrow)