*2.5. Enzyme-Linked Immunosorbent Assays*

Assays were performed as described in the ELISA kit protocols (Rat TIMP-1, Cat# RTM-100; Rat MIP3-A, Cat# DY540; Rat TNF-alpha, Cat# RTA00; R&D Systems, Minneapolis, MN, USA). Briefly, dilutions of rat plasma were added to precoated plates, incubated for 2 h at 37 ◦C. Biotin-conjugated antibody specific for the assayed protein was then added and incubated for 1 h at 37 ◦C, washed, avidin-conjugated horseradish peroxidase was added, followed by washing and TMB substrate addition. The reaction was incubated for 10–30 min, stopped with sulfuric acid and read at 450 nm.
