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. 2001 Jul 31;98(16):9110–9115. doi: 10.1073/pnas.171300598

Figure 2.

Figure 2

αS forms higher Mr aggregated species in the cytosols of mouse brain and αS-transfected MES cells. (A) The appearance of higher Mr αS immunoreactive forms is temperature-dependent. Aliquots of S370 cytosol (20 μg) from normal mouse brain were incubated at the indicated temperatures for 18 h, loaded on an 8–16% Tris-Glycine SDS gel, and blotted with H3C Ab. *, Gel-excluded material. (B) Appearance of the higher Mr αS immunoreactive forms is time-dependent. Aliquots (20 μg) of S370 cytosol from human wt αS transfected MES cells were incubated at 65°C for the indicated times and blotted with human αS-specific Ab LB509. (C) The αS-reactive higher Mr material is immunospecific. The indicated αS Abs were preabsorbed (+) or not (−) at 4°C overnight with 5 μg purified αS per microliter Ab and used to blot duplicate aliquots (20 μg) of S370 (from wt human αS-transfected MES cells) that had been incubated either at 0°C or 65°C overnight. (D) αS monomer is shifted upwards by ≈1 kDa on 18 h incubation at 65°C (also visible in C). S370 aliquots (20 μg) from wt αS-transfected MES cells were blotted with LB509.