Fig. 4

Confocal imaging of SH activity in relation to myeloperoxidase (MPO), a marker for neutrophils. Sections went through the tissue-ABPP protocol to label SHs (red) and were thereafter immunostained for MPO (yellow), followed by DAPI staining to visualize nuclei (blue). Panel a shows overall staining pattern throughout the coronal section plane. A control section undergoing identical staining protocol with no primary antibody is illustrated at top. Panel b shows staining pattern in glioma region characterized by intense SH activity originating from individual cells (TAMRA-FP hotspots). Panel c shows staining pattern in glioma region characterized by intense SH activity originating from cell clusters (TAMRA-FP hotspot clusters). Panel d shows staining pattern in healthy brain (cortex). Note detectable expression of MPO in the glioma (a). Note MPO-positive cells in the region of TAMRA-FP hotspots and marked co-localization of MPO with the TAMRA-FP signal (b). Note abundance of MPO-positive cells within TAMRA-FP hotspot clusters, as well as close match of MPO staining with TAMRA-FP hotspot clusters (c). MPO is not visible in control cortical region (d). Primary antibody rabbit anti-MPO (Abcam, cat# ab9535), dilution 1:25, secondary antibody Goat anti-rabbit IgG-Alexa Fluor 647 conjugate, dilution 1:100. Sections were from female rat 11. Scale bars: 1 mm in a, 20 μm in b-d. Images were adjusted for brightness and contrast