Paraffin sections from animals’ stomachs were deparaffinised and hydrated to distilled water. Sections were then blocked with 5% BSA solution in PBS for 1 hour at room temperature and then incubated with the appropriate primary antibody overnight at 4 °C. The primary antibody used was the anti-human Transthyretin (TTR) from DAKO (A000202) at a dilution of 1/500. The slides were then incubated with Invitrogen Alexa Fluor 555 fluorescence secondary antibody for 1hour at room temperature at 1/2,000 (anti-rabbit A-21428). Finally sections were washed with PBS and mounted using the DAKO Fluorescence Mounting Medium (S3023).
Further analysis was carried out using supplementary antibodies; Fas (Santa Cruz Biotechnology anti-rabbit sc-10231/1000) and activated Caspase-3 (Santa Cruz Biotechnology anti-goat sc-1225 1/500) to assess apoptosis, GRP78 (Santa Cruz Biotechnology anti-rabbit sc-13968 1/1000) to assess the endoplasmic reticulum stress response, and C5b-9 (EMD Millipore anti-rabbit 204903 1/4,000) to assess complement activation and Megalin (Santa Cruz Biotechnology anti-goat sc-16478 1/400). The appropriate secondary antibodies were used, anti-rabbit (Alexa Fluor A-21428 1/2000) and anti-goat (Alexa Fluor A-21432 1/2000). ECL was used to visualize the antibodies as previously explained.
Further analysis was carried out using supplementary antibodies; Fas (Santa Cruz Biotechnology anti-rabbit sc-10231/1000) and activated Caspase-3 (Santa Cruz Biotechnology anti-goat sc-1225 1/500) to assess apoptosis, GRP78 (Santa Cruz Biotechnology anti-rabbit sc-13968 1/1000) to assess the endoplasmic reticulum stress response, and C5b-9 (EMD Millipore anti-rabbit 204903 1/4,000) to assess complement activation and Megalin (Santa Cruz Biotechnology anti-goat sc-16478 1/400). The appropriate secondary antibodies were used, anti-rabbit (Alexa Fluor A-21428 1/2000) and anti-goat (Alexa Fluor A-21432 1/2000). ECL was used to visualize the antibodies as previously explained.