D-Glucosamine → L-Glutamic Acid

MION-47 and amino-CLIO (25-35 nm) were prepared as described elsewhere. D-Glucose, D-(+)-Glucosamine hydrochloride, succinic anhydride, Concanavalin A (ConA) and Sephadex G-25 were from Sigma Aldrich Co. 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) and sulfo-N-hydroxysuccinimide (sulfo-NHS) were from Pierce (Rockford, Ill.). To synthesize a glucose-functionalized nanoparticle (Glu-CLIO), NH2-CLIO was first converted to a carboxylic group functionalized nanoparticle, followed by coupling of 2-amino-glucose using a water-soluble carbodiimide. To obtain a carboxylic functionalized CLIO, 2.0 mg succinic anhydride was added into 200 uL NH2-CLIO (10 mg Fe/mL, 42 NH2 per 2064 Fe) with 300 uL (0.1 M) NaHCO3 buffer, pH 8.5. The mixture was incubated at room temperature for two hours and succinic acid removed using a Sephadex G-25 column eluted with MES buffer (0.5 M NaCl, 0.05 M MES), pH 6.0. To conjugate 2-amino-glucose to carboxy functionalized CLIO (CLIO-COOH), 2 mg EDC and 2 mg sulfo-NHS were added into 500 uL CLIO-COOH (mg Fe/mL) in MES buffer, pH 6.0. The mixture was allowed to react for two hours at room temperature and purified by Sephadex G-25 column eluted with PBS buffer at pH 7.4. Subsequently 2 mg Glucosamine was added into above solution and the mixture reacted for one hour at room temperature. Unreacted glucosamine was removed with Sephadex G-25 in PBS.

1. customsuccinic acid removed
2. washcolumn eluted with MES buffer (0.5 M NaCl, 0.05 M MES), pH 6.0
3. additionwere added into 500 uL CLIO-COOH (mg Fe/mL) in MES buffer
4. customto react for two hours at room temperature
5. custompurified by Sephadex
6. washcolumn eluted with PBS buffer at pH 7.4
7. additionSubsequently 2 mg Glucosamine was added into above solution
8. customthe mixture reacted for one hour at room temperature
9. customwas removed with Sephadex

ORDレコード

ord-3b450427d3a74af79c1ae6f05bcc4c20

出典データセット

uspto-grants-2013_09

DOI

10.6084/m9.figshare.5104873.v1

特許番号

US08535949B2