Optimizing exothermic chemical dyes within polymeric surfaces for high efficiency laser capture microdissection (LCM)

Abstract

The tumor microenvironment is a heterogeneous population of tissue and cell types that promote tumor growth. Technologies to study the tumor microenvironment typically rely on immuno-staining to identify single markers of different tissue cell subtypes without further analysis. One method to overcome these methods is Laser Capture Microdissection (LCM). LCM utilizes laser energy to melt a polymeric capture surface, or "cap", to precisely remove cells of interest from a tissue section for downstream analysis. Conventional LCM systems have an infrared (808 nm) laser. Next-generation systems contain a near-UV (405 nm) "blue" laser to obtain single-cell microdissections. To achieve high precision microdissections, a novel polymeric melting cap needed to be developed for multiple laser types. Candidate near-UV chemical dyes with absorbance near 405 nm wavelengths were dissolved within a volatile solution and mixed with a Ethylene-vinyl acetate polymeric slurry for deposition onto a cap surface body. After drying, the polymer was subjected to heat and pressure for 2 minutes followed by immediate submersion into dry ice. The cap was inspected for polymeric surface thickness, clarity, and contamination. Cap performance was measured by the AccuLift system on ovarian cancer tissue cases at various laser powers and duration settings. It was found that a combination of a near-UV absorbing dye, a photo-initiating resin, and infrared absorbing dye was required to transmit the 405 nm laser energy to melt and capture single cells (<15 micron). This next generation cap will be critical to single-cell biology research for cancer and neurodegenerative diseases such as Alzheimer’s.