BIOLUMINESCENCE BINDING ASSAY FOR BIOTIN WITH ATTOMOLE DETECTION BASED ON RECOMBINANT AEQUORIN

Biotinylated recombinant aequorin was used in the development of a hom ogeneous bioluminescence assay for biotin. It was demonstrated that th e Ca2+-triggered luminescent reaction of biotinylated aequorin can be inhibited by the presence of avidin, and this inhibition io biolumines cence intensity can be used to determine the biotin concentration in a sample. The optimized assay thus developed had a detection limit for biotin of 4 amol (equivalent to 1 X 10(-14) M in the assay system), wh ich to our knowledge is the lowest detection limit reported to date fo r a homogeneous competitive binding assay.