Quantikine rat il 1β il 1 f2 immunoassay

At sacrifice, the right lungs were injected 4 times with 3 ml aliquots of warm calcium- and magnesium-free phosphate buffered saline (PBS) (pH 7.4). The BAL fluids were then centrifuged for 7 min at 500 × g, and the BAL cells collected and re-suspended in 1 ml of PBS for evaluation. The total cell number was determined using a hemocytometer. The cells were first smeared and then stained with Wright Giemsa Sure Stain to allow counts of the total number of cells, macrophages, polymorphonuclear cells (PMNs), and lymphocytes. Two hundred cells were evaluated for the cell differentiation. Furthermore, the BAL samples were also analyzed using a blood biochemical analyzer (Hitachi 7108, Hitachi, Japan) to determine the levels of lactate dehydrogenase (LDH), micro-albumin (mALB), micro-total protein (mTP), and blood urea nitrogen (BUN). The levels of inflammatory cytokines (TNF-α, IL-1β) in the BAL fluid were measured using a Quantikine Rat IL-1β/IL-1F2 Immunoassay (R&D Systems, Inc., Minneapolis, MN) and Quantikine Rat TNF-α Immunoassay (R&D Systems, Inc., Minneapolis, MN) according to the manufacturer’s instructions (principle: Sandwich enzyme immunoassay)