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| 1 | Characterization of a new monoclonal anti-glypican-3 antibody specific to the hepatocellular carcinoma cell line, HepG2显示文摘AIM To characterize the antigen on HepG2 cell that is specifically recognized by a new monoclonal antibody raised against human liver heparan sulfate proteoglycan(HSPG), clone 1E4-1D9.METHODS The antigen recognized by m Ab 1E4-1D9 was immunoprecipitated and its amino acid sequence was analyzed LC/MS. The transmembrane domain, number of cysteine residues, and glycosylation sites were predicted from these entire sequences. Data from amino acid analysis was aligned with glypican-3(http://gffzz7fe8ccc2a30242bfs0ub5ocqk6fu56pc6.ffgz.tsg.suse.edu.cn/Tools/msa/clustalo/). The competitive reaction of mA b 1E4-1D9 and anti-glypican-3 on HepG2 cells was demonstrated by indirect immunofluorescence and analyzed by flow cytometry. Moreover, co-immunoprecipitation of mA b 1E4-1D9 and anti-glypican-3 was performed in HepG2 cells by Western immunoblotting. The recognition by mA b 1E4-1D9 of a specific epitope on solid tumor and hematopoietic cell lines was studied using indirect immunofluorescence and analyzed by flow cytometry.RESULTS Monoclonal antibody 1E4-1D9 reacted with an HSPG isolated from human liver and a band of 67 kD was detected under both reducing and non-reducing conditions. The specific antigen pulled down by m Ab 1E4-1D9, having a MW of 135 kD, was analyzed. The results showed two sequences of interest, gi30722350(1478 amino acid) and gi60219551(1378 amino acid). In both sequences no transmembrane regions were observed. Sequence number gi30722350 was 99.7% showed a match to FYCO1, a molecule involved in induction of autophagy. Sequence number gi60219551 contained 15 cysteines and 11 putative glycosylation sites with 6 predicted N-glycosylation sites. It was also matched with all PDZ domain proteins. Moreover, it showed an 85.7% match to glypican-3. Glypican-3 on HepG2 cells competitively reacted with both phycoerythrin-conjugated anti-glypican-3 and mA b 1E4-1C2 and resulted in an increase of double-stained cell population when higher concentration of m Ab 1E4-1D9 was used. Moreover, antigens precipitated from HepG2 cell by anti-glypican-3 could be detected by mA b 1E4-1D9 and vice versa. The recognition of antigens, on other solid tumor cell lines, by m Ab 1E4-1D9 was studied. The results demonstrated that m Ab 1E4-1D9 reacted with Huh7, HepG2, HT29, MCF7, SW620, Caco2, B16F1, U937, K562 and Molt4 cells. It was also found to be weakly positive to SW1353 and HL60 and negative to H460 and Hela cell lines. CONCLUSION All findings show that mA b 1E4-1D9 specifically recognizes glypican-3. Moreover, a new partner molecule of glypican-3, FYCO1 is proposed based on the results from co-precipitation studies. | Preeyanat Vongchan Robert J Linhardt | 2017 | World Journal of Hepatology2017，9，7： | 2 |
| 2 | lout resistance of conventional pedicle screw implantation in comparison to fluoroscopic computer-assisted technique显示文摘 | Linhardt O Perlick L Renkawitz T | | 0，，05： | 1 |
| 3 | Capillary electrophoresisfor the analysis of glycosaminoglycans and glycosaminoglycan-derived oligosaccharides显示文摘 | MAO W J Thanawiroon C Linhardt R J | 2002 | Biomed Chromatogr2002，16，2： | 1 |
| 4 | Vascular access and closure in coronary angiography and percutaneous intervention 显示文摘 | Byrne RA Cassese S Linhardt M | 2013 | Nat RevCardiol2013，10，1： | 1 |
| 5 | Heparin: structure and activity显示文摘 | Linhardt R J | 2003 | J Med Chem2003，46，13： | 1 |
| 6 | Ionic liquid-mediated selective extraction of lignin from wood leading to enhanced enzymatic cellulose hydrolysis显示文摘 | LEE S H DOHERTY T V LINHARDT J S | 2009 | Biotechnol Bioeng2009，102，5： | 1 |
| 7 | Lessons learned from the contamination ofheparin显示文摘 | Liu H Zhang Z Linhardt R J | 2009 | Nat Prod Rep2009，26，3： | 1 |
| 8 | Oligosaccharide mapping of Low Molecular Weight Heparins: Structure and activitydifferences显示文摘 | Linhardt RJ Loganathan D Alhakim A | 1990 | J Med Chem1990，33，： | 1 |
| 9 | Occupation-related findings in patients with degenerative disc disease of the lumbar spine:Investigation of assessment relevant questions of the occupational disease number 2108 by descriptive representation of the data from the German spinal study显示文摘 | Linhardt O Bolm-Audorff U Bergmann A | | 0，，01： | 1 |
| 10 | Purification and characterization of heparinase from Flavobacterium heparinum显示文摘 | Linhardt R J Bernstein H | 1985 | J Biological Chemistry1985，260，： | 1 |
| 11 | Purification and characterization of heparin lyases from Flavobacterium heparinum显示文摘 | Linhardt R J | 1992 | J Biological Chemistry1992，267，24： | 1 |
| 12 | Synthesis and applications of sucrose based esters 显示文摘 | Polat T Linhardt R | 2001 | J Surf Deterg2001，4，： | 1 |
| 13 | Antitumor effect of butanoy- lated heparin with low anticoagulant activity on lung cancer growth in mice and rats显示文摘 | Yu L Y Garg H G Linhardt R J | 2010 | Curr Cancer Drug Tar2010，10，： | 1 |
| 14 | Chemoenzymatic synthesis of heparan sulfate and heparin显示文摘 | Liu J Linhardt RJ | 2014 | Nat Prod Rep2014，31，12： | 1 |
| 15 | An enzymatic system for removing heparin in extracorporeal therapy显示文摘 | Linhardt R J | 1982 | Science1982，217，： | 1 |
| 16 | Increase in the growth inhibition of bovine pulmonary artery smooth muscle cells by an O- hexanoyl low-molecular-weight heparin derivative 显示文摘 | Garg H G Hales C A Linhardt R J | 2006 | Carbohydr Res2006，341，： | 1 |
| 17 | Ionic liquid -mediated selective extraction of lignin from wood leading to enhanced enzymatic cellulose hydrolysis 显示文摘 | LEE S H DOHERTY T V LINHARDT R J | 2009 | Biotechnol Bioeng2009，102，： | 1 |
| 18 | Human milk glycosaminoglycans inhibit HIV glycoprotein gp120 binding to its host cell CD4 receptor显示文摘 | Newburg DS Linhardt RJ Ampofo SA Yolken RH | 1995 | J Nutr1995，125，： | 1 |
| 19 | Structure of heparin derived tetrasaccharides显示文摘 | Merchant ZM Kin YS Linhardt RJ | 1985 | Biochem J1985，229，： | 1 |
| 20 | Exploration of the action pattern of Streptomyces hyaluronate lyase using high resolution capillary electrophoresis 显示文摘 | Park Y Cho S Linhardt RJ | 1997 | Biochim Biophys Acta1997，1337，2： | 1 |
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