Characterization of human IgG subclasses within intraocular compartments

Multiple factors regulate the distribution of the human immunoglobulin (Ig) G1-4 subclasses within different compartments of the human eye. An experimental protocol was developed for analysis of IgG1-4 using a combination of high pressure liquid chromatography (HPLC) chromatofocusing and an enzyme-linked immunosorbent assay (ELISA). IgG in the human cornea, aqueous humor, and vitreous was extracted and fractionated by chromatofocusing in a pH 9.0-6.0 gradient. A monoclonal antibody based capture-type ELISA was used to determine the IgG1-4 subclass distribution within the fractions. Histograms generated were then used to determine the IgG1-4 subclass profile for each intraocular compartment. Results indicate that the IgG species are similar but unique for each compartment. In contrast to the distribution for normal serum/plasma IgG1-4, the intraocular profiles were extremely heterogeneous and variable. These results indicate that the intraocular compartments are in a state of disequilibrium. There are apparently both common and unique factors that regulate the distribution of individual IgG subclass species for each intraocular compartment. In addition to differences in fluid dynamics, tissue biochemistry, and barrier properties, parameters, unique to the IgG subclass species may also profoundly effect intraocular localization and retention.