Computational protocol: Inhibition of SHIP2 in CD2AP-deficient podocytes ameliorates reactive oxygen species generation but aggravates apoptosis

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Protocol publication

[…] Generation of the CD2AP−/− knockout mouse model has been described earlier. Kidney samples of 3 weeks old wild type and CD2AP−/− mice were embedded in Tissue-Tek® O.C.T. Compound (Sakura, Leiden the Netherlands) and fixed with 2% paraformaldehyde (PFA) after sectioning. Sections were stained with anti-8-OHdG (15A3) with Mouse on Mouse Polymer IHC Kit (Abcam, Cambridge, UK). Slides were counterstained with haematoxylin and digitally scanned with 3DHistech Pannoramic 250 FLASH II (3DHISTECH Ltd., Budapest, Hungary). Quantification of the signal in glomeruli (n = 50–100 glomeruli per kidney) of three wild type and four CD2AP−/− mouse kidneys was performed with HistoQuant module (3DHISTECH Ltd.). [...] All cell culture experiments were carried out at least three times. All variables were presented as mean ± SD or ±SEM. The significance of differences between groups was determined by ANOVA or Students t-test (GraphPadPrism6, GraphPad Software Inc, La Jolla, CA, USA), and p-values of less than 0.05 were considered statistically significant. […]

Pipeline specifications

Software tools HistoQuant module, GraphPad Prism
Applications Miscellaneous, Whole slide imaging
Organisms Mus musculus, Homo sapiens
Chemicals Oxygen, Puromycin Aminonucleoside