THE LIMULUS BIOASSAY FOR ECDYSTEROIDS

¹ Kenyon College, Gambier, Ohio 43022

This paper presents a description of the Limulus larval bioassay for ecdysteroid compounds. The Limulus assay is routinely more sensitive than the traditional insect tests and can detect 20-hydroxyecdysone in the low picogram range when test larvae are at the point of new cuticle formation. It compares favorably with detection of ecdysteroids by radioimmune assay for routine use. Limulus larvae can detect a wide range of natural ecdysteroids and synthetic analogs and can tolerate high concentrations of ethanol or methanol solutions. Therefore, it may be a useful assay for crude alcohol extracts and tests of fractions for total ecdysteroid content. Structure-function studies show similarities and differences between Limulus and some insect systems. Premolt larvae can detect cyasterone and ponasterone A in the low picogram range, 2-deoxy-20-hydroxyecdysone in the low nanogram range. The latter compound, which is very effective in Calliphora stygia, is the least effective of the natural ecdysteroids tested in Limulus larvae.