Patent
Co-incubation of an amyloid protein with sulfated macromolecules as a method for the formation of amyloid plaques. The amyloid protein may be the beta-amyloid protein or the prion protein or the like. Amyoid plaque formation in one embodiment proceeds in vitro and desirably produces amyloid plaques that stain with Congo red and demonstrate a maltese-cross pattern when viewed under polarized light. The method also produces amyloid plaques that demonstrate an “amyloid star” appearance when viewed by transmission electron microscopy. An in vivo assay is also presented for selecting a candidate therapeutic agent for inhibiting or disrupting amyloid plaque deposition or persistence.
1. A method for the formation of particular amyloid plaques, the method comprising in vitro co-incubation of beta-amyloid protein 1–40 (SEQ ID NO: 1) for at least 3–7 days at 30–45° C. with heparan sulfate, whereby spherical or compact shaped amyloid plaques are formed that demonstrate a Maltese-cross pattern when stained with Congo red and viewed under polarized light, and an amyloid star appearance when viewed by transmission electron microscopy, and further wherein the co-incubation is at a molar ratio of beta-amyloid protein to heparan sulfate within a range of 1:0.5 to 1:100.
2. The method of claim 1 wherein the molar ratio of beta-amyloid protein to heparan sulfate is about 1:5.
3. The method of claim 1 wherein the step of co-incubation of the amyloid protein with heparan sulfate is in distilled water or Tris-buffered saline (pH 7.0–7.4).
4. The method of claim 1 wherein the step of co-incubation has a duration of about 7 days.
5. The method of claim 1 wherein the step of co-incubation of the beta-amyloid protein with heparan sulfate occurs at about 37° C.
6. A method for the formation of particular amyloid plaques, the method comprising in vitro co-incubation of beta-amyloid protein 1–40 (SEQ ID NO: 1) for at least 3–7 days at 30–45° C. with heparan sulfate, whereby spherical or compact shaped amyloid plaques are formed that demonstrate a Maltese-cross pattern when stained with Congo red and viewed under polarized light, and an amyloid star appearance when viewed by transmission electron microscopy, and further wherein the co-incubation is at a weight ratio of beta-amyloid protein to heparan sulfate within a range of 1:0.4 to 1:100.
7. The method of claim 6 wherein the weight ratio of beta-amyloid protein to heparan sulfate is about 1:8 or 1:16.
8. The method of claim 6 wherein the step of co-incubation of the amyloid protein with heparan sulfate is in distilled water or Tris-buffered saline (pH 7.0–7.4).
9. The method of claim 6 wherein the step of co-incubation has a duration of about 7 days.
10. The method of claim 6 wherein the step of co-incubation of the beta-amyloid protein with heparan sulfate occurs at about 37° C.