Establishment of a highly sensitive enzyme-linked immunosorbent assay for interleukin-1 alpha employing a fluorogenic substrate.
We have previously established a non-competitive solid-phase enzyme-linked immunosorbent assay (ELISA) specific for interleukin-1 alpha (IL-1 alpha) using a combination of polyclonal antibody as the immobilized antibody, biotinylated monoclonal antibody as the second antibody and avidin-peroxidase. The level of detection of that ELISA was 200-500 pg/ml. In order to improve its sensitivity, we have used streptavidin-beta-D-galactosidase and the fluorogenic substance 4-methylumbelliferyl-D-galactopyranoside as enzyme substrate. With this system IL-1 alpha could be detected at concentrations as low as 10-50 pg/ml, which was about 10-20 times more sensitive than conventional mouse thymocyte co-stimulator assays. Furthermore, the assay system was specific for IL-1 alpha in that neither IL-1 beta nor interleukin-2 (IL-2) interfered.
Mukaida
N
N
Department of Clinical Pathology, Jichi Medical School, Tochigi-ken, Japan.
Kasahara
T
T
Ko
Y
Y
Kawai
T
T
D000911:Antibodies, Monoclonal; D001681:Biological Assay; D001710:Biotin; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005697:Galactosides; D006801:Humans; D006923:Hymecromone / Q000031:analogs & derivatives; D007375:Interleukin-1 / Q000032:analysis*; D009598:Nitrophenylgalactosides; D011994:Recombinant Proteins / Q000032:analysis
D000911:Antibodies, Monoclonal; D005697:Galactosides; D007375:Interleukin-1; D011994:Recombinant Proteins; C055012:2-nitrophenylgalactoside; D009598:Nitrophenylgalactosides; D006923:Hymecromone; C026845:4-methylumbelliferyl-galactopyranoside; D001710:Biotin