Through the application of a simplified in vitro assay for chemotaxis of polymorphonuclear leukocytes, potent chemotaxins have been found to exist in cotton mill dust extracts. These substances may be involved in the recruitment of cells to pulmonary surfaces and in the etiology of byssinosis in textile workers who inhale cotton dust. The chemotaxins were extracted from cotton dust with alkaline aqueous solution and purified by chromatography on paper and Sephadex G-25. The active agents were readily water-soluble, stable to heat at 100 degrees C in water, inactivated by hydrolytic conditions, anionic, nonfluorescent, and had molecular weights of 200 to 2,000 daltons. Lacinilene, a fluorescent component of cotton dust reported by others to be chemotactic, was excluded by these properties and did not stimulate migration of leukocytes obtained from a variety of animal species. Pluronic polyol F68, a polypropylene glycol used as a dispersing agent, was found to represent a new class of chemokinetic substances and may account for earlier reports of chemotactic activity of lacinilene.