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. Author manuscript; available in PMC: 2015 Mar 1.
Published in final edited form as: J Immunol. 2014 Feb 3;192(5):2133–2142. doi: 10.4049/jimmunol.1301286

Figure 6. Lymphatic endothelial heparan sulfate is essential for optimal oligomerization of CCL21 and CXCL12.

Figure 6

HS was purified from cultured hLEC monolayers (grown in 10-cm plates) that were transfected with either control siRNA (siDS) or siRNA targeting the indicated HS biosynthetic enzymes, and resuspended in 500 μL of PBS. Recombinant human CCL21 (hCCL21, 20 ng/reaction, molecular weight 12.2 kDa, A) or hCXCL12 (20 ng/reaction, molecular weight: 8 kDa, B) was incubated with 0.1 μL and 0.3 μL of each HS prep, followed by BS3 mediated crosslinking, and separation on SDS-PAGE gels with detection by Western immunoblotting. Green signal: hCCL21 (A) or hCXCL12 (B) as detected by Western immunoblot; red signal: protein size marker. − and +: pure hCCL21 or hCXCL12 in the absence or presence of BS3 crosslinker, respectively.