Biomarkers such as carbon monoxide (CO) and cotinine (a nicotine metabolite) are used in tobacco cessation studies to assess smoking status. CO is easy to assess, is inexpensive, and provides immediate results. However, the short half-life of CO may limit its ability to identify smokers who have abstained for several hours. Quantitative methods (e.g., gas chromatography/mass spectrometry, or GC/MS) for measuring urine cotinine, which has a longer half-life, are valid and reliable, though costly and time consuming. Recently developed semiquantitative urine cotinine measurement techniques (i.e., urine immunoassay test strips, or ITS) address these disadvantages, though the value of ITS as a means of identifying abstaining smokers has not been evaluated. The present study examined ITS as a measure of smoking status in temporarily abstaining smokers. A total of 236 breath and urine samples were collected from smokers who participated in two separate studies involving three independent, 96-hr (i.e., Monday-Friday), Latin-square-ordered, abstinence or smoking conditions; a minimum 72-hr washout separated each condition. Each urine sample was analyzed with GC/MS and ITS. Under these study conditions, CO demonstrated moderate sensitivity (83.1%) and strong specificity (100%), whereas ITS assessment showed strong sensitivity (98.5%) and weak specificity (58.5%). In this study of short-term abstinence, ITS classified as nonabstinent nearly half of the samples collected from abstaining smokers. However, it classified nearly all nonabstinent smokers as currently smoking. Validation of ITS using GC/MS results from smokers undergoing more than 96 hr of abstinence may be valuable.