Purpose: Several small proteomic studies suggest that the prosecretory tear protein lacritin may be selectively downregulated in dry eye syndrome and in blepharitis, yet little information is available about normal baseline levels. This study assessed lacritin levels in tears from healthy individuals and addressed whether they differ according to sex, age, or time of day.
Methods: Rabbit antibodies against lacritin N-terminal peptide EDASSDSTGADPAQEAGTS (Pep Lac N-Term) were generated and characterized against human recombinant lacritin and N-65 truncation mutant. Basal tears were collected from 66 healthy individuals ranging in age from 18 to 52 years, and at four times during one 24-hour period from 34 other individuals. Lacritin levels were then analyzed by ELISA and Western blotting.
Results: Anti-Pep Lac N-Term bound lacritin, but not truncation mutant N-65 that lacks the N-terminal antigenic site. Tear lacritin levels followed a normal distribution with a mean of 4.2 ± 1.17 ng/100 ng total tear protein. Levels differed little by age or sex, and decreased slightly between 4 and 8 hours in a 24-hour cycle. Tear-blocking effects were minimal, as suggested by spiking of tears with recombinant lacritin.
Conclusions: Anti-Pep Lac N-Term-detectable lacritin comprises ~4.2 ng/100 ng total tear protein in healthy individuals, with no significant differences between males and females or among individuals between 18 and 52 years old. Levels decrease slightly in the late afternoon. These findings provide a baseline for future immunodiagnostic studies of lacritin in dry eye and other ocular diseases.