Phage antibodies

• US 6,265,150 B1
• Filed: 05/26/1998
• Issued: 07/24/2001
• Est. Priority Date: 06/07/1995
• Status: Expired due to Term

• Current Assignee: Crucell Holland B.V (Johnson & Johnson)
• Original Assignee: Becton, Dickinson & Co, Crucell Holland B.V (Johnson & Johnson)
• Inventors: Terstappen, Leon WMM, Logtenberg, Ton
• Primary Examiner(s): Bansal, Geetha P.
• Application Number:
  US09/085,072
• Time in Patent Office: 1,155 Days
• Field of Search: 435/69.6, 435/69.4, 435/332, 435/6, 435/320.1, 435/5, 530/387.1, 530/395, 436/546, 436/547
• US Class Current: 435/5
• CPC Class Codes: C07K 16/2809 against the T-cell receptor (TcR)-CD3 complex
• CPC Class Codes: C07K 16/2887 against CD20
• CPC Class Codes: C07K 2317/21 from primates, e.g. man
• CPC Class Codes: C07K 2317/622 Single chain antibody (scFv)

Abstract

Peripheral blood leucocytes incubated with a semi-synthetic phage antibody library and fluorochrome-labeled CD3 and CD20 antibodies were used to isolate human single chain Fv antibodies specific for subsets of blood leucocytes by flow cytometry. Isolated phage antibodies showed exclusive binding to the subpopulation used for selection or displayed additional binding to a restricted population of other cells in the mixture. At least two phage antibodies appeared to display hithereto unknown staining patterns of B lineage cells. This approach provides a subtractive procedure to rapidly obtain human antibodies against known and novel surface antigens in their native configuration, expressed on phenotypically defined subpopulations of cells. Importantly, this approach does not depend on immunization procedures or the necessity to repeatedly construct phage antibody libraries.

Claims

1. A method for obtaining a phage particle comprising an antibody fragment directed against an antigen associated with the surface of target cells in a heterogeneous cell population, the method comprising:

(a) providing a library of phage particles that express antibody fragments on the surface of the phage particles;

(b) incubating the library of phage particles with said target cells under conditions that allow binding of the antibody fragment expressed on the surface of the phage particles to said antigen associated with said target cells;

(c) separating said target cells and phage particles bound therewith from phage particles not bound to target cells; and

(d) recovering the phage particles bound to the target cells.

2. A method according to claim 1, wherein the separating of said target cells and phage particles bound therewith from phage particles not bound with the target cell is accomplished by flow cytometry.

3. A method according to claim 1, further comprising isolating antibody fragments which bind to said target cells.

4. A method according to claim 2, wherein the target cells are labeled with fluorochrome-labeled antibodies.

5. A method according to claim 1, further comprising repeating steps (b) through (d).

6. A method according to claim 1, wherein the library of phage particles comprises phage particles expressing Fab or single chain Fv (scFv) antibody fragments.

7. A cell-type specific library of phage particles produced according to the method of claim 1.

8. An antibody or antibody fragment obtained using the cell-type specific library of phage particles of claim 7.