The physical interaction between RANTES and cell surface proteoglycans. IPs from WCEs were performed with specific antibodies as indicated. Samples were then separated on 10% SDS-PAGE and Western blots developed using polyclonal anti-RANTES antibodies. (A) HeLa-CD4 cells were treated (+) with 640 nM RANTES for 24 hours or mock treated (–). (B) HeLa-CD4 cells were treated with RANTES as described. Following IP with indicated antibodies, GAG digestion with heparinase/chondroitinase-ABC enzymes was performed. (C) HeLa-CD4 cells were treated for 24 hours with RANTES (50 and 640 nM) or [⁴⁴AANA⁴⁷] RANTES (640 nM) or were mock treated (–). C (control), 5 μg RANTES per lane; C*: 1 μg [⁴⁴AANA⁴⁷] RANTES mutant was directly immunoprecipitated with anti-RANTES antibodies. (D) Primary CD4⁺ T cells were treated with RANTES (50 and 640 nM) for 24 hours or mock treated (–). CD44-specific IP was performed and RANTES associated with CD44 detected as described. R indicates RANTES; Rd, RANTES dimer; IgL, immunoglobulins light chain. One of 2 to 4 individual experiments is shown.