Quarterly Report, July - September 1989
Abstract
The first quarter has been devoted in part to organization, ordering, and reagent preparation. We have ordered, obtained, and are in the process of setting up the computer work station, the beta scintillation counter and the low pressure chromatography system. As part of the work plan we have made killed whole cell vaccines to E. coli 0111B4 and S. typhimurium, and generated polyclonal rabbit antisera to each that have titers of 1:3000,000 by ELISA. We are now purifying the IgG on DEAE sephacel anion exchange columns, and we are this week coupling S. typhimurium LPS to sepharose beads in order to affinity- purify the IgG. If the procedure goes without difficulty, we will couple the E. coli 0111B4 LPS in the next several weeks also. We have ordered rabbits for the preparation of large pools of normal and tolerant sera to use as constant beginning material for the purification. We have also started to set up the cytotoxicity assay for tumor necrosis factor using 929 cells, a task made easier by a modern cell culture facility in the new laboratory. Our early results suggest that a murine macrophage cell line (RAW 264.7) will serve as a macrophage source (alleviating the cumbersome need for rabbit peritoneal cells), and our preliminary results with these cells indicate that the assay is sensitive to picograms/ml E. coli 0111B4 LPS. We will try this assay, in addition to limulus lysate, as an on line check for the neutralizing ability of our preparations during the purification.
Document Details
- Document Type
- Technical Report
- Publication Date
- Oct 10, 1989
- Accession Number
- ADA213396
Entities
People
- H. S. Warren
Organizations
- Massachusetts General Hospital