Development of an enzyme immunoassay for arginine-vasopressin (AVP)-like insect diuretic hormone.
1. The AVP-like insect diuretic hormone is a biologically active antiparallel dimer present, along with its non-active monomeric form (Cys-Leu-Ile-Thr-Asn-Cys-Pro-Arg-GlyNH2), in the African locust. 2. It exhibits diuretic activity by increasing fluid excretion at the level of the Malpighian tubules. 3. To date, both monomer and dimer have been assayed using a radioimmunoassay originally prepared for mammalian AVP. 4. We have developed here an original enzyme immunoassay based on the use of antibodies to insect AVP-like raised in rabbits against synthetic monomers and dimers, using acetylcholinesterase conjugate as an enzymatic tracer. 5. This enzyme immunoassay enables measurement of the dimer to be made with adequate sensitivity (0.3 nmol/l, i.e. 21 pg/well) and reproducibility while sensitivity of the monomer is somewhat lower (14 nmol/l, i.e. 480 pg/well). 6. The assay was validated by assaying native dimer and monomer throughout the different steps of purification (from a crude extract to reversed-phase liquid chromatographic fractions). 7. A good correlation was observed between radioimmunoassays and enzyme immunoassays. 8. The enzyme immunoassay was also used to measure the level of AVP-like peptides in several insect tissues not explored to date.
Proux
J
J
Laboratoire de Neuroendocrinologie, URA CNRS 1138, Université de Bordeaux 1, Talence, France.
Baskali
A
A
Remy
C
C
Creminon
C
C
Pradelles
P
P
D000595:Amino Acid Sequence; D000818:Animals; D001127:Arginine Vasopressin / Q000032:analysis*; D004231:Diuresis / Q000502:physiology*; D006110:Grasshoppers / Q000737:chemistry*; D007124:Immunoenzyme Techniques; D007301:Insect Hormones / Q000032:analysis*; D008297:Male; D008969:Molecular Sequence Data; D011863:Radioimmunoassay; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D007301:Insect Hormones; D001127:Arginine Vasopressin