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. Author manuscript; available in PMC: 2016 Nov 1.
Published in final edited form as: Alzheimers Dement. 2015 Apr 4;11(11):1286–1305. doi: 10.1016/j.jalz.2015.01.005

Fig. 5. Aqueous extracts of human brain contain up to 4 different sized Aβ assemblies separable by SEC.

Fig. 5

TBS extract (1 ml) from one AD brain (AD33) was chromatographed on a Superdex 75 10/30 HR column (A and B). Half ml fractions were collected, a 0.25 ml aliquot was lyophilized and analyzed by WB using a combination of 2G3 and 21F12 (A) or a monoclonal antibody, 46-4, which does not recognize Aβ (B). Bands specifically reactive with anti-Aβ mAbs were detected in fxs 1 and 2, (High mwt), 3, and 4 (Int mwt) 17 and 18 (monomer). A darker exposure (A inset) is shown to highlight bands in fxs 17 and 18. Two ml of AD-TBS (AD28) was chromatographed on a Superdex 75 10/30 HR column linked in tandem with a Superdex 200 10/30 HR column (C). Fractions (1 ml) were lyophilized and analyzed by WB using 2G3/21F12. Because the fractions from a single chromatographic run couldn’t be accommodated on a single gel, samples were electrophoresed on 2 (A and B) or 3 (C) gels alongside synthetic Aβ1-42 standards (not shown) which were used to normalise the exposure of each blot. Species that migrated on SDS-PAGE as monomer and dimer are indicated by arrows and molecular weight markers shown on the left, and fraction numbers are provided below each lane. Four peaks of Aβ immunoreactivity were detected in the SEC fractions, and based on their elution relative to SEC standards are designated as (i) high molecular weight (high mwt), (ii) intermediate molecular weight (Int mwt), (iii) ~7 kDa Aβ species and (iv) true monomer (monomer) - these species are highlighted with an underline. The elution of linear dextran standards is indicated by up-pointing arrows and labelled 5, 12, 25, 80, 150, 270 kDa, while the elution of globular standards, is indicated by down-pointing arrows labelled 1, 17, 44, 158 and 670 kDa. Fraction 0 indicates the peak fraction in which Blue dextran eluted. Non-specific indicates bands detected when 46-4 was used.