Primary rat hippocampal neurons show lewy body inclusion formation and loss of cells when treated with Type 1 mouse Alpha Synuclein Protein Pre-formed Fibrils (SPR-324) at 4 µg/ml (D-F) on DVI2, but not when treated with a control (A-C). Tissue: Primary hippocampal neurons. Species: Sprague-Dawley rat. Fixation: 3% formaldehyde from PFA for 20 min. Blocker: 1:1 PBS:LiCOR Odyssey Block (LiCOR, 927-40010) and 30 mL/mL of 0.1% triton-X 100 for 30 min. Primary Antibody: Mouse anti-pSer129 Antibody (1:1000) and Rabbit anti-pSer129 (1:800) for 24 hours at 4°C. Secondary Antibody: ATTO 546 Donkey Anti-Mouse (1:700) and ATTO 488 Donkey Anti-Rabbit (1:700) for 1 hour at RT (composite green). Counterstain: Hoechst (blue) nuclear stain at 1:3000 for 1 hour at RT. Localization: Lewy body inclusions. Magnification: 20x.
C57/BL6 mice were injected with sonicated recombinant mouse alpha synuclein monomers or fibrils at 8 weeks of age. Mice were unilaterally injected in the dorsal striatum (bregma AP + 0.2 mm, L +/1 2.0 mm, V - 3.0 mm) and sacrificed 30 days post-injection. (A) 1.25 uL mouse alpha synuclein monomers (SPR-323). (B) 2.5 uL mouse alpha synuclein monomers (SPR-323). (C) 2.5 ug alpha synuclein PFFs (SPR-324). (D) 5 ug alpha synuclein PFFs (SPR-324) Inset: PBS (negative control). Primary antibody: Anti-Alpha Synuclein pSer129 (SMC-600) at 1:10 000. Secondary antibody: anti-rabbit HRP. Mice injected with PFF displayed alpha synuclein staining in the striatum and cortex and contralateral to the injection site. Courtesy of: Porsolt.
C57/BL6 mice were injected with 5 ug sonicated mouse recombinant alpha synuclein PFFs (SPR-324) at 8 weeks of age. Mice were unilaterally injected in the dorsal striatum (bregma AP + 0.2 mm, L +/1 2.0 mm, V - 3.0 mm) and sacrificed 30 days post-injection. (A) contralateral cortex. (B) ipsilateral cortex. (C) contralateral striatum. (D) ipsilateral striatum. Primary antibody: Anti-Alpha Synuclein pSer129 (SMC-600) at 1:10 000. Secondary antibody: anti-rabbit HRP. Mice injected with PFF displayed alpha synuclein staining in the striatum and cortex and contralateral to the injection site. Courtesy of: Porsolt.
Immunohistochemistry analysis of rat brain injected with Type 1 mouse alpha synuclein PFFs (SPR-324). Species: Female Sprague-Dawley Rat. Rat was injected with 16µg Type 1 mouse alpha synuclein PFFs (SPR-324) in each of 2 injection sites: AP+1.6, ML+2.4, DV-4.2 from skull; and AP-1.4, ML+0.2, DV-2.8 from skull. 30 days post-injection. Fixation: Saline perfusion followed by 4% PFA fixation for 48 hrs. Primary antibody: rabbit monoclonal anti-pSer129 alpha synuclein. Secondary Antibody: Biotin-SP Donkey Anti-Rabbit IgG (H+L) at 1:500 for 2 hours in cold room with shaking. ABC signal amplification, DAB staining. Magnification: 20X. Alpha synuclein pathology is seen in the periform/insular cortex and the cingulate cortex on both the same (ipsi) and opposite (contra) sides as the injection sites.
Primary mouse hippocampal neurons treated with 100 nM sonicated mouse alpha synuclein PFFs (SPR-324) (A). Phosphorylated alpha synuclein (detected with pSer129 antibody SPC-742) was visible in perinucleus and neurites compared to untreated control (B). Read the protocol at pabmabs.com/?p=7901. Image courtesy of Trine Rasmussen, Simon Molgaard Jensen at Aarhus University.
Supplier Page from StressMarq Biosciences Inc for Alpha Synuclein Pre-formed Fibrils