In this Phase II application Dr. Boraker proposes to continue his studies enhancing ELISA performance using AM technology. The rate-limiting factor in solid-phase immunoassays (such as the enzyme-linked immunosorbent assay or ELISA) is diffusion. When a cylindrical polystyrene probe is immersed in the microwell contents and caused to oscillate to and fro in a transverse linear fashion, a stable pattern of 8 high-velocity vortices appears in the well (called """"""""acoustic microstreaming,"""""""" or); the rapid liquid motion produced by these vortices reduces diffusion layer thickness and thereby enhances transport of macromolecules onto the microwell surface. During the Phase I study, it was observed that the acoustic probe itself, although only l9 percent of the surface area, produced 60-90 percent of the signal of unstirred control wells. The investigators propose, therefore, to optimize acoustic probe ELISA by studying the independent and interdependent variables of probe shape, diameter, frequency and amplitude to maximize diffusion rate. They then propose to build l) a manual AM instrument capable of enhancing ELISA instrument which would process the equivalent of 5 microplates simultaneously, and 2) a prototype automated ELISA instrument which would process the equivalent of 5 microplates by performing all steps from sample incubation to chromophore development. Advantages of this innovation are: reduced reliance on individual well timing, simplified, rapid probe washing; """"""""batch"""""""" processing in conjugate; retention of original sample; elimination of the """"""""quenching"""""""" step; ease of servicing and cleaning, and reduced cost. Such an automated instrument could have wide appeal to diagnostic companies as world-wide demand for ELISA automation continues to drive the nonistopic immunoassay marketplace.
| Boraker, D K; Bugbee, S J; Reed, B A (1992) Acoustic probe-based ELISA. J Immunol Methods 155:91-4 |
