Visualization is crucial to the effective analysis of biological pathways. A poorly laid out pathway confuses the user, while a well laid out one improves the user's comprehension of the underlying biological phenomenon. We present a new, elegant algorithm for layout of biological signaling pathways. Our algorithm uses a force-directed layout scheme, taking into account directional and rectangular regional constraints enforced by different molecular interaction types and subcellular locations in a cell. The algorithm has been successfully implemented as part of a pathway visualization and analysis toolkit named Patika, and results with respect to computational complexity and quality of the layout have been found satisfactory. The algorithm may be easily adapted to be used in other applications with similar conventions and constraints as well. Patika version 1.0 beta is available upon request at http://www.patika.org. © 2004 Elsevier Inc. All rights reserved.