Cervical lymph nodes were sonicated in a 50 mM Tris-HCl, 150 mM NaCl, and 1% Triton-X, pH 7.4 buffer containing a complete protease inhibitor cocktail tablet (Roche Applied Science, Mannheim, Germany). Subsequently, samples were centrifuged at 10,000g for 20 minutes at 4°C to obtain organ supernatants. For multiplex analysis, 25 μL of the lymph node supernatants were employed using the MILLIPLEX MAP mouse cytokine/chemokine panel (MCYTMAG-70K-PX32; Millipore Corporation, Billerica, MA, USA) and MAGPIX Multiplexing Instrument (Millipore) according to the manufacturer's instructions. Eight analytes were studied: IL-2, IL-4, IL-5, IL-13, IFN-γ, TNF-α, eotaxin, and regulated upon activation normal T cell expressed and presumably secreted (RANTES). The concentration of analytes was determined by MAGPIX Xponent software (Millipore), and the results are reported as the mean ± SEM.