Protein Components of Amyloid

Abstract

AMYLOID may be deposited in animal or human connective tissues in a variety of pathological circumstances. It consists mainly of protein, with up to 5 per cent of carbohydrates largely in the form of mucopoly-saccharides. Previous attempts to characterize the protein moiety of amyloid have included : (a) Extraction with strong alkali followed by electrophoresis, revealing either α-1-, β-2-, and β-globulins with traces of albumin¹ : or α-globulin alone² ; (b) Extraction with strong alkali followed by analytical ultra-centrifugation, demonstrating a single component with a sedimentation coefficient of 6² ; (c) Extraction with concentrated aqueous urea followed by analytical ultracentrifugation, showing five fractions with sedimentation coefficients of 1, 6, 9, 16 and 23 (ref. 3); (d) Extraction with strong alkali followed by amino-acid analysis, which gave no precise protein identification but excluded collagen as a significant constituent on the basis of low hydroxyproline values²; (e) Fluorescent protein tracing in histological preparations, which is so sensitive a technique that trace contaminants would be revealed, has detected the presence of γ-globulin^4–6, β-1 complement⁶, rheumatoid factor⁶, and fibrinogen⁶; (ƒ) Absorption of specific anti γ-globulin antiserum with purified amyloid substance, which produced no significant reduction in titre⁷; (g) Extraction with concentrated aqueous urea followed by immuno-diffusion and electrophoresis against specific antisera to albumin, γ-globulin, cæruloplasmin, orosomucoid, β-2 macroglobulin, rheumatoid factor, C-reactive protein, a pool of myeloma globulins, and whole serum ; none of these antisera precipitated the amyloid proteins³; (h) Extraction with strong alkali followed by preparation of an antiserum which, on immuno -electrophoresis against whole serum, gave precipitin arcs in the α-2 regions⁸. None of these methods has entirely surmounted the two main obstacles to precise identification of the protein components of amyloid. These are, first, the relative insolubility of this material in all but chemically vigorous reagents which are liable to degrade proteins and, secondly, the difficulty in obtaining amyloid free from incidental contamination by plasma proteins. The new method reported here appears to circumvent both these difficulties in the isolation of pure natural amyloid.