Isolation of antibodies which neutralise the activity of early pregnancy factor.
Early pregnancy factor (EPF) is a secreted protein with growth regulatory and immunomodulatory properties. It functions as an autocrine growth factor for tumour cells and as an autocrine or paracrine growth factor for regenerating normal cells. Anti-EPF antibodies have demonstrable anti-tumour activity and, as a result, hybridomas which produce such antibodies are unstable. In this study, the phage display antibody techniques have been investigated as a means of producing recombinant anti-EPF antibodies. Mice were immunised with synthetic peptides which correspond to the N or C terminal regions of EPF, and their splenic tissue was used to make combinatorial antibody libraries. The Fab repertoire was displayed on the surface of phage and panned over recombinant EPF. Reactive Fabs were identified by ELISA and their binding was characterised by BIAcore analysis and functional studies. Three libraries with a size of greater than 5x10(7)cfu were constructed and a total of 26 unique Fabs with specific reactivity against EPF were identified. Three Fabs were purified and of these one demonstrated strong EPF neutralising activity, one had intermediate activity and the other was not neutralising. Phage display has provided the means of circumventing the problems of anti-EPF hybridoma development and has resulted in the production of antibodies with potential applications in the diagnosis of pregnancy and the diagnosis and therapy of cancer.
Hammond
F
F
Department of Medical Oncology, St. Vincent's Hospital, Victoria St., Darlinghurst, NSW 2010, Sydney, Australia.
Cavanagh
A
A
Morton
H
H
Hillyard
N
N
Papaioannou
A
A
Clark
M
M
Wanigesekera
D
D
Swanton
M
M
Ward
R
R
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D000918:Antibody Specificity; D018835:Chaperonin 10; D003429:Cross Reactions; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D019151:Peptide Library; D010455:Peptides / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D011257:Pregnancy Proteins; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D013491:Suppressor Factors, Immunologic
D000906:Antibodies; D018835:Chaperonin 10; D019151:Peptide Library; D010455:Peptides; D011257:Pregnancy Proteins; D011994:Recombinant Proteins; D013491:Suppressor Factors, Immunologic; C025124:early pregnancy factor