Recognition of adenocarcinoma in situ (AIS) in cervical cytology is challenging. We calculated the sensitivity and accuracy of Papanicolaou (Pap) tests obtained within 1 year of a histologic diagnosis of AIS from 2007 to 2016. We also correlated it with the coexistence of squamous lesions, calculated the interobserver agreement, and compared these measures with those of endocervical adenocarcinoma (ECCA). We correlated AIS detection with high-risk human papillomavirus (hrHPV) status. Of 72 patients with histologic AIS and 48 patients with ECCA, 92% and 87.5%, respectively, had abnormal Pap test results. A glandular abnormality was detected in 44.4% of the AIS and 77.1% of the ECCA cases. Complete cytohistologic concordance was reached in 8.3% of AIS and 22.9% of ECCA cases. In addition, 27.8% of AIS and 6.3% of ECCA cases were diagnosed on Pap as a high-risk squamous abnormality. Concurrent squamous lesions were present in 79.2% of patients with AIS and 29.2% of patients with ECCA. The Paps from the AIS and ECCA cases were diagnosed as pure squamous abnormalities in 47.2% and 10.4% of cases, respectively. In the AIS cases, interobserver agreement was substantial for detection of any high-risk cytologic abnormality (kappa = 0.67) and fair for detection of any glandular abnormality (kappa = 0.34). Among the 26 patients with AIS tested for hrHPV, 92% had positive results and 8% had negative results. The cytologic sensitivity for the detection of AIS remains low. It is directly related to the coexistence of squamous lesions. Cytology and hrHPV as stand-alone screening tests fail in the early detection of a small proportion of glandular lesions, although combined testing will improve their detection rates. Copyright © 2019 American Society of Cytopathology. Published by Elsevier Inc. All rights reserved.