Detecting biomolecules in picoliter vials using aequorin bioluminescence

The quantitative determination of proteins in picoliter-volume vials is described. The assay is based on the bioluminescence of the photoprotein aequorin along with photon-counting detection. Using this approach, avidin can be detected at femtomole levels by taking advantage of its inhibitory effect on the bioluminescence signal generated by biotinylated recombinant aequorin. The picoliter vials were fabricated on glass substrates using a laser ablation technique. Parameters that affect the reproducibility of the assay such as the fabrication and calibration of the pipets, the fabrication of the vials, and the composition of the assay solutions were studied.