Application of a filamentous phage pVIII fusion protein system suitable for efficient production, screening, and mutagenesis of F(ab) antibody fragments.
We describe the application of a novel filamentous phage vector system suitable for efficient screening and production of F(ab) antibody fragments. The vector system can concurrently produce free F(ab) fragments and F(ab) displayed on the surface of M13 bacteriophage via a VHCH1-pVIII fusion protein. When expressed in a supO (nonsuppressor) strain of Escherichia coli free F(ab) can be produced. Antibody F(ab) fragments are secreted into culture medium at concentrations up to 0.3 mg/liter and conveniently subjected to detailed analysis with little or no purification. Higher concentrations of F(ab) (approximately 10 mg/liter) were found to accumulate in the periplasmic space. In this report the vector system is shown to produce correctly folded and assembled F(ab) fragments of chimeric L6, a mAb against a tumor-associated Ag expressed by many human carcinomas.
Huse
W D
WD
Ixsys, Inc., San Diego, CA 92121.
Stinchcombe
T J
TJ
Glaser
S M
SM
Starr
L
L
MacLean
M
M
Hellström
K E
KE
Hellström
I
I
Yelton
D E
DE
D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000302:isolation & purification; D017104:Bacteriophage M13 / Q000378:metabolism*; D001483:Base Sequence; D002213:Capsid / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000276:immunology; D005822:Genetic Vectors; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D016296:Mutagenesis; D014760:Viral Fusion Proteins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D014760:Viral Fusion Proteins