Design and peptide-based validation of phage display antibodies for proteomic biochips.
To validate potential application of phage display-antibody arrays for high-throughput screening on a novel proteomics biochip, we examined the epitopes versus the full protein of glucose-6-phosphate-dehydrogenase (G6PD) from yeast. In a predictive approach, we used the Hopp-Woods method and compared the results with antibodies directed against the entire enzyme. In total, 16 peptides of a length of 11 amino acids each fulfilling the desired criteria were identified and synthesized. Subsequently, antibodies against G6PD were raised using a phage display library. Selective interaction of the antibodies with certain peptides facilitated the identification of epitopes predicted by the hydropathic profile. The setup was adapted to a novel biochip system based on surface-enhanced absorption for direct CCD-camera based screening.
Stich
N
N
Kluyver Laboratory for biotechnology, TU-Delft, Julianalaan 67, 2628BC Delft, The Netherlands.
van Steen
G
G
Schalkhammer
T
T
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D000915:Antibody Affinity; D000918:Antibody Specificity; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping / Q000379:methods; D005954:Glucosephosphate Dehydrogenase / Q000737:chemistry / Q000276:immunology; D018625:Microscopy, Atomic Force; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019151:Peptide Library; D040081:Protein Array Analysis / Q000379:methods*; D040901:Proteomics / Q000379:methods*; D012441:Saccharomyces cerevisiae / Q000201:enzymology
D000906:Antibodies; D019151:Peptide Library; D005954:Glucosephosphate Dehydrogenase