Enhanced chemiluminescence labeled-antibody immunoassay (Amerlite-MAB) for free thyroxine: design, development, and technical validation.
We describe the development and validation of a one-step nonradioactive immunoassay for free thyroxine (Amerlite-MAB FT4) in serum or plasma, in the dedicated Amerlite enhanced luminescence assay system. A monoclonal antibody to thyroxine (T4), conjugated with horseradish peroxidase (HRP; EC 1.11.1.7), competes for binding with FT4 or with a conjugate of a protein and L-triiodothyronine immobilized onto the surface of the assay microwells. The signal is generated by antibody-HRP bound to the protein-T3 conjugate, with luminol as substrate. The assay design exploits the high sensitivity of luminescence signal detection, permitting minimal sample dilution and T4 sampling by the antibody. It withstands progressive dilution of serum and is unaffected by T4-binding proteins in serum. The disclosure and validation of this FT4 assay follows guidelines recommended by the American Thyroid Association.
Christofides
N D
ND
Cardiff Laboratories, Kodak Clinical Diagnostics Ltd., Amersham, UK.
Sheehan
C P
CP
D000911:Antibodies, Monoclonal; D004492:Edetic Acid; D005230:Fatty Acids, Nonesterified / Q000097:blood; D006801:Humans; D007124:Immunoenzyme Techniques / Q000706:statistics & numerical data; D007700:Kinetics; D008163:Luminescent Measurements; D010949:Plasma / Q000737:chemistry; D011228:Prealbumin / Q000378:metabolism; D011786:Quality Control; D012680:Sensitivity and Specificity; D012709:Serum Albumin / Q000378:metabolism; D013974:Thyroxine / Q000097:blood*; D013975:Thyroxine-Binding Proteins / Q000378:metabolism
D000911:Antibodies, Monoclonal; D005230:Fatty Acids, Nonesterified; D011228:Prealbumin; D012709:Serum Albumin; D013975:Thyroxine-Binding Proteins; D004492:Edetic Acid; D013974:Thyroxine