A study to evaluate the protection provided by permethrin-treated fabric following cold-water washing against biting by mosquitoes is reported. Australian Defense Force (ADF) disruptive pattern combat uniform (DPCU) shirt fabric and entire shirts were treated by dipping in a 0.6% emulsion (Perigen Defense, containing 500 g/liter permethrin), and commercial factory treatment in the United States (Factory A) and Europe (Factory B). Protection was recorded after 1, 3, 5, 10, 30, and 50 washes. The treated fabric provided 100% protection against bites of Anopheles farauti Laveran for at least 50 washes, although only 4.8–19.0% of this species fed through untreated DPCU. The protection provided by each type of permethrin treatment against Aedes aegypti (L.) biting was variable; however, there were no significant differences between the percentage of mosquitoes biting between 1 and 10 washes. A comparison between the two factory treatments for 1–50 washes also showed no statistical difference in Ae. aegypti feeding. Chemical analysis of fabric was conducted using gas chromatography and showed that the initial dose was 0.125 mg/cm² for Perigen-treated fabric, which fell to 0.004 mg/cm² after 10 washes. By contrast, factory treatments resulted in initial dose rates of 0.20 mg/cm² for Factory A and 0.19 mg/cm² for Factory B. After 10 washes, Factory A-treated fabric had 0.09 mg/cm² and Factory B 0.15 mg/cm² of permethrin. Despite the higher concentrations of permethrin in the fabric, there was not a commensurate increase in biting protection provided by the factory-treated fabric, compared with fabric treated by dipping in permethrin emulsion.