This study was performed to investigate the inflammatory changes occurring in the human conjunctiva at different time periods after allergen provocation. Twenty-three ryegrass-sensitive patients with allergic conjunctivitis (19 with hayfever and four with vernal conjunctivitis) were challenged by topical administration of ryegrass antigen to the eye. Allergen concentrations were increased in increments until an immediate ocular allergic reaction was elicited. Numbers of various inflammatory cells (neutrophils, eosinophils, lymphocytes, and monocytes) found in conjunctival scrapings were quantified and correlated with the clinical profile, total serum IgE, and serum IgE to Rye I antigen. Twenty minutes after some level of antigen topical challenge to the eye, all patients had ocular redness, tearing, and itching. Compared with findings in seven control subjects, significant inflammatory cells were found in the conjunctival scrapings of patients before challenge (p less than 0.05) and 20 minutes (p less than 0.001) and 6 hours (p less than 0.002) after effective challenge. Significant increases in neutrophils of patients occurred after 20 minutes (p less than 0.001), and in eosinophils at 6 hours (p less than 0.005), compared with values of control subjects. When each case was evaluated individually, nine of the 23 patients had highly evident inflammatory changes 6 hours after allergen provocation. The levels of total serum IgE and serum IgE to Rye I antigen of these nine patients did not differ significantly from the other patients in the study. Our data provide the first evidence in humans that significant inflammatory changes in conjunctival scrapings are present long after allergen exposure has ended.