Since their invention in 1986 [1], optical tweezers have become increasingly valuable tools for research in the biological [2] and physical [3] sciences. Using a focused beam of light to trap and move matter, optical tweezers offer convenient, non-invasive access to processes at the mesoscopic scale. Most applications, however, have involved manipulating small numbers of particles or small volumes of soft materials because existing optical tweezer implementations can create just a few tweezers at once. Were they readily available, large arrays of optical tweezers could be used to organize microscopic particles into complex structures, to sort them intelligently, to study collective behavior in many-body systems, and to manipulate materials too delicate to trap with a single tweezer. We recently described [4] a method to create arrays of optical tweezers using computer-generated holographic beam splitters. This Article further explains how to design and fabricate the necessary holograms and how to integrate them into holographic optical tweezer arrays capable of trapping hundreds of particles simultaneously.