220.127.116.11. Solution of radioactive JH (RIA tracer solution)
The stability of JH-III in aqueous solution is limited. As this solution should be used within 4-6 weeks, it is necessary to estimate the volume needed for each assay series. 100 µl of this solution is required per sample, and the radioactivity should be 6,000-6,500 cpm/100 µl tritiated juvenile hormone III ([10-3H(N)] Juvenile hormone III, Perkin Elmer NET586050UC).
1. Tritiated juvenile hormone III comes as 50 µCi in 0.5 ml toluene/hexane and an appropriate volume to be used in an assay series must first be evaporated to dryness in a glass test tube.
2. Re-dissolve the dry radioactive JH in the appropriate volume of phosphate buffer (see section 18.104.22.168).
3 .Transfer to a screw cap glass vial.
4. Keep vial on orbital shaker for 24 h at 4oC.
5. Test radioactivity by liquid scintillation counting of 100 µl aliquots.
6. The level of radioactivity should be 6,000-6,500 cpm/100 µl.
If necessary adjust by adding phosphate buffer or tracer.