Development of novel whole-cell immunoadsorbents by yeast surface display of the IgG-binding domain.
The ZZ domain derived from Staphylococcus aureus, which binds to the Fc part of immunoglobulin G (IgG), was displayed on the cell surfaces of yeast Saccharomyces cerevisiae by cell-surface engineering using the C-terminal half of alpha-agglutinin under control of the 5'-upstream region of the isocitrate lyase gene from Candida tropicalis (UPR-ICL). Display of ZZ on the cell surface was confirmed by immunofluorescence microscopy. Enzyme-linked immunosorbent assay (ELISA) and sandwich ELISA using the S. cerevisiae cells displaying ZZ detected IgG and antigen (human serum albumin) down to a concentration of 1-10 ng/ml in both cases. The detection range covered by these assay systems was wide and could be varied by adjusting the amount of cells and reaction times with horseradish peroxidase (HRP) substrate. Moreover, yeast cells displaying ZZ were successfully used for repeated affinity purification of IgG from serum. These results indicate that S. cerevisiae displaying ZZ may constitute novel and genetically renewable whole-cell immunoadsorbents widely applicable to immunoassays and affinity purification.
Nakamura
Y
Y
Department of Chemical Science and Engineering, Faculty of Engineering, Kobe University, Japan.
Shibasaki
S
S
Ueda
M
M
Tanaka
A
A
Fukuda
H
H
Kondo
A
A
D000818:Animals; D001666:Binding Sites, Antibody; D004797:Enzyme-Linked Immunosorbent Assay; D007141:Immunoglobulin Fc Fragments / Q000378:metabolism; D007074:Immunoglobulin G / Q000032:analysis* / Q000302:isolation & purification / Q000378:metabolism; D007163:Immunosorbent Techniques; D011817:Rabbits; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D012709:Serum Albumin / Q000032:analysis; D013211:Staphylococcus aureus / Q000378:metabolism
D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D012709:Serum Albumin