Signal versus noise

Why is this important? It is relatively easy to generate a signal from an aptamer or an antibody binding to a target molecule and we have a host of options as to how this event can result in a signal. A common diagnostic trick is to immobilize a fixed amount of the target molecule. Aptamers can be readily labeled with a wide variety of signal moieties including fluorophores, gold nanoparticles (GNPs), quantum dots, or directly to colour reducing enzymes like horseradish peroxidase (HRP) through the synthesis of the aptamer with a biotin on one end or the other. Such labeled aptamers can then be mixed with your sample containing an unknown amount of the target molecule. This combined sample/aptamer mixture is then exposed to the immobilized target molecule. The higher the concentration of the target molecule in the sample the less aptamer will bind to the immobilized target. The sample is removed, the surface is washed, and the signal is measured.

Another classic approaches to signal production involve sandwich assays where a pair of aptamers are used, one is immobilized on the surface (the capture aptamer) and the other is labeled and added free in solution (detection aptamer) combined with the sample. The capture aptamer than captures the sample as well along with the detection aptamer, and the signal is observed from the detection aptamer.

More complex approaches involving the release of a signal upon binding of the aptamer to a target where the expression of the label is quenched in the absence of binding and liberated by the binding also dominate publications. Such applications can involve an allosteric shift on the part of the aptamer as a function of binding, or the displacement of a quenching antisense.

The point of this blog is how the expression of a signal affects the signal/noise ratio. With aptamers versus antibodies this can be manifested in many different ways including the binding strength of the aptamer for the target (the stronger the binding the less signal is lost in each wash step), the specificity of the aptamer for the target and the amount of signal per binding event. I will devote subsequent blogs to each of these steps and how we have overcome intrinsic aptamer difficulties with practical innovative solutions. The key always though is increasing signal per binding event while decreasing or maintaining background noise. This is achievable with aptamers, but the solutions require different approaches and different chemistries than what works with antibodies.

In the end we want to convert the scream to a symphony or at least a pop song.