Purification of LPS Binding Substances in Inflammatory Serum
Abstract
We radiolabeled E. Coli 018, E. Coli 0113, E. Coli 0111:B4 and, S. typhimurium by growing the organisms in tritated broth and extracting the 3H-LPS by the hot phenol method as described in the original grant protocol. We also prepared and aliquotted large quantities of normal and tolerant (acute phase) rabbit sera. We prepared affinity purified anti-LPS IgG by coupling LPS to epoxy-linked sepharose beads, and in turn coupled the anti-LPS IgG to sepharose beads using cyanogen bromide to prepare columns capable of affinity purifying LPS (and whatever is bound to it) from normal and tolerant sera. In later experiments we have utilized a murine Mab directed to the O-polysaccharide antigen of E. Coli 0111:B4 to prepare similar affinity columns. We incubated unlabeled and/or labeled LPS in each sera, affinity purified the complexes, eluted them with KSCN, and then analyzed the eluted material on SDS polyacrylamide gels.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jul 31, 1990
- Accession Number
- ADA225376
Entities
People
- H. S. Warren
Organizations
- Massachusetts General Hospital