Detection of pro-inflammatory cytokines in healthy canine tears using Canine Cytokine SpikeMix™ mass spectrometry via multiple reaction monitoring.

PURPOSE:To evaluate the efficacy of the Canine Cytokine SpikeMix™ and MRM-MS for detecting pro-inflammatory cytokines in canine tears from healthy research Beagles. METHODS:A complete ophthalmic examination was performed on 15 healthy research Beagles to verify no ophthalmic diseases. Tears were collected OU by placing a Weck-Cel® cellulose spear in the ventral conjunctival fornix for 1 min. The Weck-Cel® spear was placed in a 2.0 mL tube with a centrifuge filter forcing tears to flow through the filter into the bottom of the tube. The tears were analyzed using the Canine Cytokine SpikeMix™ and MRM-MS. Descriptive data from this study was reported as the normalized total peak area (nTPA) and median (range) using data imported from the online MRM-MS Skyline program. RESULTS:The level of 16 pro-inflammatory cytokines was successfully detected in all 15 dogs. The four cytokines with the highest median amounts in the samples were IL-2 = 0.1243 (0.019-6.7289), IL-6 = 0.964 (0.0036-16.9365), TNFα = 0.1644 (0.0096-0.7138), and CSF-2 = 0.4022 (0.1475-2.6208). CONCLUSIONS:This study revealed that 16 pro-inflammatory cytokines in canine tears from healthy dogs can be detected with Canine Cytokine SpikeMix™ and MRM-MS.