In assembly line positions and other occupations, employees experience pain and discomfort associated with long periods of standing. Modifying flooring is a common intervention method used to alleviate problems associated with constrained standing. This study investigated the effects of four different standing conditions upon assembly workers' perception of general fatigue and discomfort associated with various body parts following 1 week of 8-h days of exposure to each condition. The four standing conditions were (1) on hard floor, (2) on a floor mat, (3) wearing shoe in-soles, and (4) wearing shoe in-soles while standing on a floor mat. Questions were asked regarding general fatigue and discomfort. In general, the mat, in-soles and combined conditions were more comfortable than standing on hard floor. Moderately strong correlations were found for the variables of height, age and job tenure. Shorter individuals experienced greater discomfort in the upper leg and low back when standing on a mat and when wearing in-soles. The older workers and those with more job seniority reported greater discomfort associated with body joints when standing on the hard floor and when standing on the floor mat. No significant differences in fatigue or discomfort were found when comparing the overall effects of using the floor mat to wearing the shoe in-soles or the combined condition.