Abstract
Antibody phage display technology is well established and widely used
for selecting specific antibodies against desired targets. Using
conventional manual methods, it is laborious to perform multiple
selections with different antigens simultaneously. Furthermore, manual
screening of the positive clones requires much effort. The authors
describe optimized and automated procedures of these processes using a
magnetic bead processor for the selection and a robotic station for the
screening step. Both steps are performed in a 96-well microplate format.
In addition, adopting the antibody phage display technology to
automated platform polyethylene glycol precipitation of the enriched
phage pool was unnecessary. For screening, an enzyme-linked
immunosorbent assay protocol suitable for a robotic station was
developed. This system was set up using human γ-globulin as a model
antigen to select antibodies from a VTT naive human single-chain
antibody (scFv) library. In total, 161 γ-globulin-selected clones were
screened, and according to fingerprinting analysis, 9 of the 13 analyzed
clones were different. The system was further tested using testosterone
bovine serum albumin (BSA) and β-estradiol-BSA as antigens with the
same library. In total, 1536 clones were screened from 4 rounds of
selection with both antigens, and 29 different testosterone-BSA and 23
β-estradiol-BSA binding clones were found and verified by sequencing.
This automated antibody phage display procedure increases the throughput
of generating wide panels of target-binding antibody candidates and
allows the selection and screening of antibodies against several
different targets in parallel with high efficiency.
Original language | English |
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Pages (from-to) | 282-293 |
Number of pages | 12 |
Journal | Journal of Biomolecular Screening |
Volume | 14 |
Issue number | 3 |
DOIs | |
Publication status | Published - 2009 |
MoE publication type | A1 Journal article-refereed |
Keywords
- antibody phage display
- automation
- high throughput
- magnetic beads
- robotic station