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A simplified microassay for inhibition of protein synthesis in reticulocyte lysates by immunotoxins

Authors
Journal
Immunology Letters
0165-2478
Publisher
Elsevier
Publication Date
Volume
14
Issue
1
Identifiers
DOI: 10.1016/0165-2478(86)90017-9
Keywords
  • Immunotoxin
  • Protein Synthesis
  • Reticulocyte Lysate
  • Ricin
  • Toxin
Disciplines
  • Biology

Abstract

Abstract We describe a simplified cell free reticulocyte lysate protein synthesis assay which has been shown to reliably demonstrate dose dependent inhibition of protein synthesis with cycloheximide and with four ricin A chain immunotoxin conjugates. Maximal [ 3H]leucine incorporation was observed using 40 μl of reticulocyte lysate per microtiter well and by incubating samples for 30 minutes before harvesting with a multiple automated sample harvester. Trichloroacetic acid precipitation and hydrogen peroxide decolorization were not required with this method. This rapid, simplified assay system will facilitate testing of agents that inhibit protein synthesis, particularly immunotoxins.

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