An enzyme immunoassay for synthetic thymulin.
Thymulin, a metallononapeptide with the following aminoacid sequence: pyroGlu-Ala-Lys-Ser-Gln-Gly-Gly-Ser-AsnOH is a thymic hormone involved in T cell differentiation requiring zinc to express biological activity as measured by the rosette assay. We established an enzyme immunoassay (EIA) for synthetic zinc-free thymulin with a thymulin-acetylcholinesterase conjugate as tracer and specific polyclonal rabbit antithymulin antibodies. The assay is performed as a classical competition assay in microtiter plates previously coated with mouse monoclonal IgG to rabbit IgG. A quantitative thymulin assay more sensitive than radioimmunoassays (RIAs) previously described was obtained with a sensitivity (IC50) of 32.5 +/- 5 pg/ml and a detection limit of 5 pg/ml. Analysis in the EIA of synthetic thymulin analogs showed that the minimal peptidic structure necessary for enzymatic tracer competition is the C-terminal part Lys3 to Asn9. It was also shown that the biologically active form of thymulin (zinc-bound) has the same immunoreactivity as zinc-free thymulin and that other thymic hormones, thymosin alpha 1 and thymopoietin II (or TP5) and unrelated short peptides do not cross-react with thymulin. These data demonstrate the specificity of this EIA for thymulin and show its suitability for application in biological fluids.
Métreau
E
E
INSERM U 25, Hôpital Necker, Paris, France.
Pléau
J M
JM
Dardenne
M
M
Bach
J F
JF
Pradelles
P
P
D000918:Antibody Specificity; D003429:Cross Reactions; D006728:Hormones / Q000032:analysis; D007124:Immunoenzyme Techniques; D011485:Protein Binding; D013935:Thymic Factor, Circulating / Q000032:analysis* / Q000378:metabolism; D013951:Thymus Hormones / Q000032:analysis* / Q000276:immunology; D015032:Zinc / Q000378:metabolism
D006728:Hormones; D013951:Thymus Hormones; D013935:Thymic Factor, Circulating; D015032:Zinc