Lymphatic vessels form a critical component in the regulation of human health and disease. While their functional significance is increasingly being recognized, the comprehensive heterogeneity of lymphatics remains uncharacterized. Here, we report the profiling of 33,000 lymphatic endothelial cells (LECs) in human lymph nodes (LNs) by single-cell RNA sequencing. Unbiased clustering revealed six major types of human LECs. LECs lining the subcapsular sinus (SCS) of LNs abundantly expressed neutrophil chemoattractants, whereas LECs lining the medullary sinus (MS) expressed a C-type lectin CD209. Binding of a carbohydrate Lewis X (CD15) to CD209 mediated neutrophil binding to the MS. The neutrophil-selective homing by MS LECs may retain neutrophils in the LN medulla and allow lymph-borne pathogens to clear, preventing their spread through LNs in humans. Our study provides a comprehensive characterization of LEC heterogeneity and unveils a previously undefined role for medullary LECs in human immunity. Display omitted •Single-cell RNA sequencing of human lymph nodes unveils six types of LECs•LECs lining the floor and ceiling of the SCS, MS, and valve are the main types•LECs of the SCS floor and MS highly express neutrophil chemoattractants•Human MS LECs support neutrophil adhesion in the LN medulla via CD209 Using single-cell RNA sequencing, Takeda and colleagues identify six types of lymphatic endothelial cells (LECs) in human lymph nodes (LNs) mapped to particular locations, including subcapsular and medullary sinuses (SCSs and MSs). CD209 on MS LECs mediates neutrophil adhesion in the human LN medulla, which may prevent pathogen spread through LNs.