The discovery of luminol electrogenerated chemiluminescence (ECL) at carbon electrodes in the presence of hydrogen peroxide by Sakura in 1992 and the enhanced ECL of ruthenium-(2,2′-bipyridyl)32+ complex (Ru(bpy)32+) in the presence of reduced nicotinamide adenine dinucleotide (NADH) by Jameison in 1996 opened the way for ECL-based enzyme assays: the activity of oxidases that generate hydrogen peroxide and the one of dehydrogenases that produce NADH could be measured thanks to light emission of luminol or Ru(bpy)32+, respectively. This chapter reviews the important achievements in ECL-based biosensing systems since nearly 20 years. Numerous systems involving, first luminol or Ru(bpy)32+ as soluble ECL emitters, then immobilized on an electrode surface are described, distinguishing bioassays and biosensors. The introduction of nanomaterials such as nanoparticles (NPs) that act as ECL enhancers with some of them having their own ECL properties (e.g. quantum dots) are presented. The combination of several nanomaterials into the so-called nanocomposites offers new opportunities in the field of enzyme-based ECL bioassays and biosensors. Finally, future trends in bipolar electrochemistry and ratiometric assays are presented.