The use of ethanol for preserving proteins and glycoproteins of the trabecular meshwork

We evaluated methods for storage of the trabecular meshwork and iris-ciliary body for protein and glycoprotein analysis. The trabecular meshwork and iris-ciliary body of rabbit eyes were microdissected and stored in Laemmli sample buffer, Carnoy’s fluid (75% ethanol-25% glacial acetic acid), or 100% ethanol at ambient temperature, 4°C, -20°C, or -80°C for 24 h or 30 days. Fresh and stored tissues were processed for one-dimensional polyacrylamide gel electrophoresis (PAGE) and Western blot using Con A lectin. The protein patterns of stored and fresh tissues as determined by silver-stained polyacrylamide gels were similar. However, ethanol-stored tissues revealed other proteins (MW of 15-30 kD and 150 kD), and the staining intensity and band resolution of lower MW (15-40 kD) were enhanced. The glycosylation patterns of stored and fresh tissues as determined by Con A (recognizes certain N-linked glycoproteins containing mannose and glucose) were also similar, but the ethanol-stored tissues stained more intensely, especially the high (>200 kD) and low (<35 kD) MW ranges. These PAGE results indicate that ethanol storage is useful for preserving and resolving the protein/glycoprotein profiles of the trabecular meshwork.