Chimeric protein for selective cell attachment onto cellulosic substrates.
We have developed a fusion protein (CBD-LG) incorporating a cellulose-binding domain and an antibody binding domain, protein LG, to provide an adaptor molecule for cell separation with regenerated cellulose hollow fiber arrays. A single hollow fiber cell adhesion assay utilizing a CD34+ cell line, KG1a, was used to investigate whether ligand affinity interactions were strong enough for cell attachment and separation. CBD-LG efficiently captured CD34+ cells labeled with the mouse IgG2a monoclonal antibody MHCD3400. However, it was not possible to bind CD34+ cells labeled with an IgG1 antibody (HPCA-2). The low affinity of HPCA-2 for LG was overcome by secondary antibodies: KG1a cells that were dual labeled with HPCA-2 followed by rat anti-mouse IgG1 adhered inside hollow fibers coated with CBD-LG. Alternatively, immobilized rabbit polyclonal anti-mouse IgG1 captured cells labeled with HPCA-2. The development of an adaptor molecule to display recombinant domains at the surface of hollow fibers will be an effective tool to investigate cellular ligand-receptor interactions, a necessary step in the development of hollow fiber bioreactors for manufacture of human cellular products.
Craig
Scott J
SJ
Graduate School of Biomedical Engineering, University of New South Wales, Australia.
Shu
Alan
A
Xu
Yin
Y
Foong
Frances C
FC
Nordon
Robert
R
D000818:Animals; D000906:Antibodies / Q000737:chemistry*; D018952:Antigens, CD34 / Q000737:chemistry; D002448:Cell Adhesion; D002469:Cell Separation; D002482:Cellulose / Q000737:chemistry*; D003001:Cloning, Molecular; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry; D008024:Ligands; D051379:Mice; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011817:Rabbits; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D013379:Substrate Specificity
D000906:Antibodies; D018952:Antigens, CD34; C522136:CBD-LG fusion protein; D007074:Immunoglobulin G; D008024:Ligands; D011993:Recombinant Fusion Proteins; D002482:Cellulose