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SH-SY5Y cells (~2.4 × 106) were spin-inoculated with MOIs of 10, 50, and 100 with each of the following: P. gingivalis [American Type Culture Collection (ATCC) BAA-308] [wild type (WT)], P. gingivalis KgPΔIg-B, and P. gingivalis ΔK/ΔRAB-A. Uninfected SH-SY5Y cells were used as a control. SH-SY5Y cells and P. gingivalis strains were centrifuged at 1000g for 10 min at RT in Dulbecco’s modified Eagle’s medium (DMEM)/F12 supplemented with 2 mM l-glutamine and BSA (200 μg/ml), followed by incubation for 1, 4, and 8 hours, respectively, in a CO2 incubator. After the indicated incubation times, cells were collected and cell pellets were lysed with 250 μl of radioimmunoprecipitation assay buffer supplemented with protease inhibitor cocktail for 10 to 15 min. Total protein (16 μg) was used for WB. WBs were probed with tau monoclonal antibody TAU-5 (Thermo-MA5-12808). Glyceraldehyde-phosphate dehydrogenase (GAPDH) was used as an internal reference.

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