C1 MCK2 is an inactive chemokine.

(A) HEK293 cells were transfected with pFuse vectors expressing human IgG-Fc fusion proteins of wt, C1 and 129stop MCK2. 24 hours after transfection, supernatants from transfected cells were harvested, proteins precipitated with EtOH and detected by WB using an antibody specific for human IgG. Supernatants from mock-transfected cells were used as a control. (B) The protein preparations from (A) were also analyzed by WB using an antibody specific for MCK2 (anti-MCK2₁₃₁). (A) and (B) The specific bands of MCK2-Fc fusion proteins are indicated by arrows. Molecular weights of marker proteins are indicated on the left. (C) Transwell migration assays were performed with J774 cells using supernatants from HEK293 cells transfected with pFuse vectors expressing the indicated MCK2 fusion proteins. The Fc fusion protein amounts were adjusted using an Fc-specific ELISA. Migration is expressed as percentage of migration of human IgG-Fc expressed from HEK293 cells transfected with the empty pFuse cloning vector. Symbols represent independent experiments. Shown are means +/- SD of independent experiments (n = 3–9). P values were determined using a one-way ANOVA test.