• Although it may seem that difference of minutes may have an impact on the assay precision, MiCo BioMed has demonstrated with multiple assays that well optimized assays on Optimiser™ easily achieve CV < 6-10%. The minimal effect of flow rate on precision is a combination of multiple factors:
    • On the micro-scale reaction kinetics are vastly different compared to the macro-scale kinetics of conventional 96-ell ELISA plate. In microfluidic channels, most surface binding reactions are saturated in ∼ 5 minutes. Optimiser™ characterization data shows that up to ∼ 75% of peak absorption is completed in only 10 seconds and assay binding reactions saturate in ∼ 5 minutes. This is a result of two factors:
      • The diffusion distances in the microchannel are extremely small (the channel has a cross-section of only 200 μm x 200 μm) hence diffusion is no longer a limiting factor.
      • The surface area of the microchannel is ∼ 1.5 times the surface area at the base of a conventional 96-well ELISA plate. The volume contained in the microchannel is ∼ 5 μl leading to ∼ 50x higher surface area to volume ratio which allows for extremely efficient binding reactions.
    • Even for the well that drains in 8 minutes, the initial section of the microchannel (towards the center) is filled up in ∼ 2-3 minutes. Optimiser™ characterization data shows that the first few loops of the microchannel contribute ∼ 95% of the optical signal hence even if the last 1-2 loops take significantly longer to fill, their contribution to the signal is almost negligible. Consequently, variations in signal from the last loops have little impact on overall (assay) signal variation.
    • For most reaction steps in the assay sequence (except for sample/standard loading step) the biomolecules are present in vast abundance and the binding reactions are completed extremely quickly. To ensure good precision, it IS recommended that the sample/standard incubation should be 20 min.
    • Finally, the incubation interval (when there is no liquid left in the well) “smooths out” the effect of flow rate variations.