This paper discusses the effect of mass (chemical quantity) of injected dose on positron emission tomography (PET) and single-photon emission computed tomography (SPECT). Commonly, PET or SPECT imaging study uses a "no-carrier added" dose, which contains a small amount of radioactive imaging agent (in picogram to microgram). For small animal (rodent) imaging studies, specifically targeting binding sites or biological processes, the mass (chemical quantity) in the dose may significantly modify the binding, pharmacokinetics and, ultimately, the imaging outcome. Due to differences in size and other physiological factors between humans and rodents, there is a dramatic divergence of mass effect between small animal and human imaging study. In small animal imaging studies, the mass, or effective dose (ED(50)), a dose required for 50% of receptor or binding site occupancy, is usually not directly related to binding potential (B(max)/K(d)) (measured by in vitro binding assay). It is likely that dynamic interplays between specific and nonspecific binding in blood circulation, transient lung retention, kidney excretion, liver-gallbladder flow, soft tissue retention as well as metabolism could each play a significant role in determining the concentration of the tracer in the target regions. When using small animal imaging for studying drug occupancy (either by a pretreatment, coinjection or chasing dose), the mass effects on imaging outcome are important factors for consideration.