Figure - PMC

Skip to main content

View full-text article in PMC

. Author manuscript; available in PMC: 2006 May 31.

Published in final edited form as: Cancer Res. 2005 Jul 1;65(13):5750–5760. doi: 10.1158/0008-5472.CAN-04-4653

ESL-1 is a Candidate E-selectin Ligand on Human Bone-Metastatic Prostate Tumor Cells. Membrane protein and immunoprecipitates were resolved on 4-20% SDS-PAGE gels, transferred to PVDF membrane and blotted as indicated. In Panel A, ESL-1 was identified from the gel fragment (box) corresponding to the E-selectin-Ig-reactive membrane protein at 150kDa from MDA PCa 2b cells by mass spectrometry. Immunoblot analysis of (+) control WEHI-3 and of MDA PCa 2b membrane protein (40μg/Lane) with ESL-1 anti-sera showed a stainable protein at 150kDa. To determine whether ESL-1 was the E-selectin glycoprotein ligand at 150kDa, E-selectin-Ig affinity-purified eluate from MDA PCa 2b membrane protein (300μg) was blotted with ESL-1 anti-sera (Panel B). Also, anti-ESL-1 immunoprecipitates of MDA PCa 2b membrane protein (300μg) were immunoblotted with E-selectin-Ig (Panel C). In Panel B, the 150kDa protein was detected in E-selectin-Ig eluate with ESL-1 anti-sera and not in the E-selectin-Ig eluate containing EDTA. In Panel C, the 150kDa protein was blotted with E-selectin-Ig in membrane protein (40μg/Lane) and anti-ESL-1 immunoprecipitates from WEHI-3 and MDA PCa 2b cells. Immunoblots performed with AP-secondary Ab alone did not result in background staining. Immunoblots are representative of a minimum of 5 experiments.

ESL-1 is a Candidate E-selectin Ligand on Human Bone-Metastatic Prostate Tumor Cells. Membrane protein and immunoprecipitates were resolved on 4-20% SDS-PAGE gels, transferred to PVDF membrane and blotted as indicated. In Panel A, ESL-1 was identified from the gel fragment (box) corresponding to the E-selectin-Ig-reactive membrane protein at 150kDa from MDA PCa 2b cells by mass spectrometry. Immunoblot analysis of (+) control WEHI-3 and of MDA PCa 2b membrane protein (40μg/Lane) with ESL-1 anti-sera showed a stainable protein at 150kDa. To determine whether ESL-1 was the E-selectin glycoprotein ligand at 150kDa, E-selectin-Ig affinity-purified eluate from MDA PCa 2b membrane protein (300μg) was blotted with ESL-1 anti-sera (Panel B). Also, anti-ESL-1 immunoprecipitates of MDA PCa 2b membrane protein (300μg) were immunoblotted with E-selectin-Ig (Panel C). In Panel B, the 150kDa protein was detected in E-selectin-Ig eluate with ESL-1 anti-sera and not in the E-selectin-Ig eluate containing EDTA. In Panel C, the 150kDa protein was blotted with E-selectin-Ig in membrane protein (40μg/Lane) and anti-ESL-1 immunoprecipitates from WEHI-3 and MDA PCa 2b cells. Immunoblots performed with AP-secondary Ab alone did not result in background staining. Immunoblots are representative of a minimum of 5 experiments.