Vitamin B12 (B12) deficiency, although treatable, impacts up to 43% of community-living older adults; long-term care (LTC) residents may be at greater risk. Recommendations for screening require further evidence on prevalence and incidence in LTC. Small, ungeneralizable samples provide a limited perspective on these issues. The purposes of this study were to report prevalence of B12 deficiency at admission to LTC, incidence 1 year post-admission, and identify subgroups with differential risk. This multi-site (8), retrospective prevalence study used random proportionate sampling of resident charts (n = 412). Data at admission extracted included demographics, B12 status, B12 supplementation, medications, diagnoses, functional independence, cognitive performance, and nutrition. Prevalence at admission of B12 deficiency (<156 pmol/L) was 13.8%; 47.6% had normal B12 (>300 pmol/L). One year post-admission incidence was 4%. Better B12 status was significantly associated with supplementation use prior to LTC admission. Other characteristics were not associated with status. This work provides a better estimate of B12 deficiency prevalence than previously available for LTC, upon which to base protocols and policy. Prospective studies are needed to establish treatment efficacy and effect on health related outcomes.