Endocervical adenocarcinoma in situ (AIS) is not always identified on cervical Papanicolaou (Pap) test cytology because the Pap test has relatively low sensitivity for the diagnosis endocervical glandular lesions. We performed a retrospective study to determine the relative sensitivity of different diagnostic approaches, including Pap tests, cervical biopsy and/or endocervical curettage, loop electrosurgical excision procedure (LEEP), and hysterectomy specimens. Cases of endocervical AIS diagnosed from August 2005 to January 2019 were retrieved from our institution's pathology databases, and their clinicopathologic features were reviewed. A total of 74 patients with endocervical AIS with or without concurrent squamous intraepithelial lesions or cervical neoplasms were identified. Their mean age at diagnosis was 39.9 years. More than one half of the cases of AIS were not detected from screening Pap tests but were diagnosed during histologic examination of cervical biopsy or endocervical curettage, LEEP, or cone biopsy specimens (~66%). Only a few patients had had a definitive diagnosis of AIS from the Pap tests (10.8%). Other abnormal glandular cytology included atypical glandular cells, not otherwise specified (16.2%), atypical glandular cells favoring neoplasia (5.4%), and atypical glandular cells suspicious for malignancy (1.3%). Abnormal squamous cytology was common in the study population (54%), with high-grade squamous intraepithelial lesion the most common finding (30%). AIS was diagnosed in 31 of 42 cervical biopsies or curettages, with 16 cases an incidental finding and 15 cases confirming previous abnormal glandular cytology. In addition, AIS was identified in 51 of 53 LEEPs. Approximately 41.5% of those undergoing LEEP had a previous diagnosis of AIS, and 54.7% of the cases were incidental findings. More than one half of the AIS cases harbored significant concurrent cervical lesions, including 26.7% with high-grade squamous intraepithelial lesion, 5.7% with low-grade squamous intraepithelial lesion, 1.9% with invasive squamous cell carcinoma, 20.9% with invasive adenocarcinoma, and 6.7% with microinvasive adenocarcinoma. Our results have demonstrated that the ability to detect AIS with routine screening Pap testing or biopsy/curettage has variable efficacy depending on the screening methods. Given the relatively low combined sensitivity of Pap testing and biopsy/endocervical curettage in the diagnosis of AIS, all LEEPs and cervical cone biopsies performed for squamous cell abnormalities should be thoroughly evaluated for glandular lesions.