Widely diverse biological queries are now routinely analyzed on the various optical platforms: laser line scanners, nonconfocal imagers and confocal imagers. These analyses may be performed to query a limited number of samples or range to include the evaluation of a million samples as is the goal of many screening departments in the pharmaceutical drug discovery area. First we review the key elements that distinguish the optical pathway and the hardware features used amongst the three classifications of automated imaging platforms. Recognizing the need for both resolution and throughput and maximizing the use of optics we discuss some of the influences that address how to best match some of the more common biological assays with the current hardware imaging platforms. We describe here some considerations with respect to the biology, the cell types, and the goals of the screening efforts.