Linked Life Data

21772990

Source:http://linkedlifedata.com/resource/pubmed/id/21772990

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:dateCreated
2011-7-20
pubmed:abstractText
Conjugation of siRNA to macromolecules such as serum albumin has multiple potential benefits, including enhanced extravasation via albumin-mediated transcytosis across endothelial cells and reduced renal clearance. In attempting to conjugate siRNA to albumin, we used commercially sourced amine-modified siRNA and reacted it with the heterobifunctional linker succinimidyl 4-[N-maleimidomethyl]cyclohexane-1-carboxylate (SMCC) to introduce a maleimide group suitable for conjugation to the thiol group of the surface-exposed cysteine residue (Cys 34) within albumin. We found the conjugation of the SMCC-treated siRNA to bovine serum albumin (BSA) to be very inefficient and investigated the cause of the low yield of conjugate. Ultrafiltration with phosphate-buffered saline prior to activation with SMCC dramatically increased the yield of siRNA-albumin conjugate (~15-fold). Communication with the commercial supplier revealed that ammonium acetate buffer was used in a desalting step as part of the siRNA purification process prior to supply, likely resulting in ammonium counterions to the siRNA polyanion, which would interfere with conjugation by consuming the SMCC. After ultrafiltration, a greatly reduced amount of SMCC could be used to affect conjugation, without significant reduction in yield. These data indicate that amine-modified siRNA sourced commercially may require ultrafiltration or dialysis prior to use in conjugation reactions.
pubmed:commentsCorrections
pubmed:language
eng
pubmed:journal
pubmed:status
PubMed-not-MEDLINE
pubmed:issn
2090-021X
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
2011
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
154609
pubmed:year
2011
pubmed:articleTitle
Commercially Supplied Amine-Modified siRNAs May Require Ultrafiltration prior to Conjugation with Amine-Reactive Compounds.
pubmed:affiliation
Monash Institute of Pharmaceutical Sciences, Monash University, 381 Royal Parade, Parkville, VIC 3052, Australia.
pubmed:publicationType
Journal Article