1,2-Dichloroethane → L-Aspartic Acid

Conjugates of BMAA with BSA or with HRP were prepared by activating, in the absence of BMAA, the carboxylic groups of BSA or HRP with EDC thereby forming BSA-EDC or HRP-EDC. This was followed by the coupling of the activated carboxylate groups on the carrier protein to the primary amino group of BMAA. The procedure was as follows: 50 mg of EDC and 5 mg of sulfo-NHS (Sulfo-N-hydroxysuccinimide) were added to 15 mg of BSA or HRP pre-dissolved in 5 ml of 10 mM KH2PO4, pH 5.0. The reaction mixture was stirred for 2-3 minutes, thereby forming BSA-EDC or HRP-EDC, followed by the addition of 5 mL of a 1% solution of BMAA in 0.2 M K2HPO4, pH 8.5. After the solution was stirred overnight, the conjugate (BSA-EDC-BMAA or HRP-EDC-BMAA) was dialyzed exhaustively against PBS-7.4 (0.14 M NaCl, 10 mM K2HPO4, pH 7.4) or TBS-8 (tris-buffered saline, pH 8). Conjugates were stored at ≦20° C. (20° C. or less) By this procedure the conjugate was formed by linkage of BMAA to the carrier protein predominantly via the glutamic and aspartic acid residues of the carrier protein.

1. stirringAfter the solution was stirred overnight
2. customthe conjugate (BSA-EDC-BMAA or HRP-EDC-BMAA) was dialyzed exhaustively against PBS-7.4 (0.14 M NaCl, 10 mM K2HPO4, pH 7.4) or TBS-8 (tris-buffered saline, pH 8)
3. customwere stored at ≦20° C.
4. custom(20° C. or less)

ORD-Eintrag

ord-081eacc0d5f947e5a4ca457b505093bc

Quelldatensatz

uspto-grants-2013_03

DOI

10.6084/m9.figshare.5104873.v1

Patentnummer

US08394596B1