For the Clathrin and AP2 immunoprecipitation (IP) assays, protein samples were added to Dynabeads-Protein G (30 μg/100 μL) according to the manufacturer protocol (Invitrogen, DK).
The following were used for both WB and IP analysis: rabbit monoclonal (Y188) antibody to APP (ab32136, Abcam, UK), mouse anti-alpha adaptin antibody (AP6) specific against AP2 (MA1-064, Thermo Fisher, DK), mouse anti-Clathrin heavy chain antibody (clone X22) (MA1-065, Thermo Fisher, DK). Rabbit, anti-AP1+2 antibody (ab21981) were provided by Abcam.
APP pTyr residues were immunoprecipitated using anti-pTyr antibody clone 4G10® agarose conjugate (clone 16-10, Millipore) and analyzed with rabbit anti-APP (clone Y188) following the same procedures previously reported (Matrone et al., 2011 (link)). Rabbit anti-pan Fyn (#4023) and anti-Src pTyr416 (#2101) and anti-Src pTyr527 (#2105) were provided by Cell Signaling Technology (BioNordika, DK).
The following were used for both WB and IP analysis: rabbit monoclonal (Y188) antibody to APP (ab32136, Abcam, UK), mouse anti-alpha adaptin antibody (AP6) specific against AP2 (MA1-064, Thermo Fisher, DK), mouse anti-Clathrin heavy chain antibody (clone X22) (MA1-065, Thermo Fisher, DK). Rabbit, anti-AP1+2 antibody (ab21981) were provided by Abcam.
APP pTyr residues were immunoprecipitated using anti-pTyr antibody clone 4G10® agarose conjugate (clone 16-10, Millipore) and analyzed with rabbit anti-APP (clone Y188) following the same procedures previously reported (Matrone et al., 2011 (link)). Rabbit anti-pan Fyn (#4023) and anti-Src pTyr416 (#2101) and anti-Src pTyr527 (#2105) were provided by Cell Signaling Technology (BioNordika, DK).
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