Abstract:
:Affinity maturation is one of the cardinal strategies for improving antibody function using in vitro evolutionary methods; one such well-established method is phage display. To minimise gene deletion, we previously developed an open sandwich (OS) method wherein selection was performed using only phage-displaying VH fragments after mixing with soluble VL fragments. The decrease in anti-EGFR antibody 528 affinity through humanization was successfully recovered by selecting VH mutants using this OS method. However, the affinity was not similar to that of parental 528. For further affinity maturation, we aimed to isolate VL mutants that act in synergy with VH mutants. However, the OS method could not be applied for selecting VL fragments because the preparation of soluble VH fragments was hampered by their instability and insolubility. Therefore, we initially designed a modified OS method based on domain-swapping of VH fragments, from added soluble Fv fragments to phage-displaying VL fragments. Using this novel Fv-added OS selection method, we successfully isolated VL mutants, and one of the Fv comprising VH and VL mutants showed affinity almost equivalent to that of parental 528. This method is applicable for engineering other VL fragments for affinity maturation.
journal_name
Sci Repjournal_title
Scientific reportsauthors
Sanada H,Kobayashi K,Oyama K,Maru T,Nakanishi T,Umetsu M,Asano R,Kumagai Idoi
10.1038/s41598-018-23796-3subject
Has Abstractpub_date
2018-04-03 00:00:00pages
5414issue
1issn
2045-2322pii
10.1038/s41598-018-23796-3journal_volume
8pub_type
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