Abstract

DO.22.04

Comparison of Antibody Patterns in Sera and Aqueous Humor of Glaucoma Patients and Healthy Subjects by Means of Protein Microarrays

Nils Böhm, Uta Thiel, Ulrike Lossbrandt, Norbert Pfeiffer, Franz H. Grus

Experimentelle Ophthalmologie, Universitäts-Augenklinik Mainz

Objective
In the past decade, multiple studies demonstrated changes in antibody profiles against ocular antigens in sera and aqueous humor of glaucoma patients. However, none of these studies directly compared serum versus aqueous humor antibody patterns in the same patients. Using protein-microarrays we analyzed antibody levels in both for intraindividual comparison. Additionally, we compared antibody reactivities from glaucoma patients and healthy subjects.
Methods
Sera and aqueous humor of patients with primary open-angle glaucoma (POAG; n=13) and healthy controls (CTRL; n=13) were used for antibody analysis. The protein arrays were prepared by spotting 40 different purified antigens onto nitrocellulose-coated slides. The arrays were incubated with sera and aqueous humor respectively. For visualization of the antibody-antigen-reactions arrays were treated with a fluorescence labeled anti-human IgG antibody, followed by fluorescence scanning. The signals emitted from secondary antibodies were digitized and the spot intensities were compared using multivariate statistical techniques.
Results
The intraindividual comparison revealed consensuses but also differences between antibody patterns of sera and aqueous humor. In both, aqueous humor and serum, POAG patients showed more than twofold increased reactivities for α-1-Antitrypsin and Annexin V compared to healthy subjects (P≤0.001). In contrast, β-L-Crystallin showed a significantly increased reactivity in aqueous humor (POAG: ME=5049±1638; CTRL: ME=2119±673; P≤0.01) and a decreased reactivity in sera (P≤0.01) of POAG patients. Using a biomarker panel of ten antibodies/antigens from each body fluid respectively, we were able to differentiate between POAG and CTRL with a specificity and sensitivity of approx. 90% (ROC-curve; serum: r=0.91; aqueous humor: r=0.93).
Conclusions

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