Office Action 1: Non-Final Rejection (2024-09-16)
Rejection types: 103 - Obviousness

CLAIMS:
WO 2021/023619 PCT/EP2020/071533 18 CLAIMS

1\. A method for purifying an antibody comprising:
a) Loading a mixture comprising the antibody to be purified onto an affinity
chromatography resin, wherein the affinity chromatography resin is not a nickel-based, zinc-based or cobalt- based resin,
b) Washing the chromatographic resin with a wash buffer;
c) Eluting the antibody with an elution buffer that comprises 0.01 M to 1.OM imidazole or
an imidazole-analogue and a pH of 3 to 5.

2\. A method according to claim 1, wherein said affinity chromatography resin is not a metal- ion-based resin.

3\. A method according to claim 1 or 2, wherein said affinity chromatography resin is selected from: a protein A chromatography resin, a protein G chromatography resin and a protein L chromatography resin.

4\. A method according to any preceding claim, wherein the elution buffer comprises 0.01M to 0.5M imidazole or imidazole-analogue.

5\. A method according to any preceding claim, wherein the wash buffer comprises imidazole or an imidazole-analogue.

6\. A method according to claim 5, wherein the wash buffer comprises 0.01M to 0.5M imidazole or an imidazole-analogue.

7\. A method according to any preceding claim, wherein the method comprises a further step of equilibrating the chromatographic resin with an equilibration buffer comprising imidazole or an imidazole-analogue prior to loading of the mixture comprising the antibody onto the chromatographic resin.

8\. A method according to any preceding claim wherein one, two of all of, of steps a), b) and c) are performed under dynamic conditions.

9\. A method according to any preceding claim wherein the antibody is selected from: IgG, Fab', F(ab')2, scFv, Fab-Fv, Fab-scFv, Fab-(scFv)2, Fab-(Fv)2, diabodies, triabodies, and
tetrabodies. 5 10 15 20 25 30 35

OFFICE ACTION:
Notice of Pre-AIA  or AIA  Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Claims 10-18 are pending and are under examination.
Information Disclosure Statement
The information disclosure statement (IDS) submitted on 2/1/2022 and 8/5/2024  has been considered and an initialed copy is enclosed.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA  35 U.S.C. 102 and 103 (or as subject to pre-AIA  35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA  to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary.  Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claim(s) 10-13 and 17-18  is/are rejected under 35 U.S.C. 103 as being unpatentable over Heywood et al. US 2018/0100007 4/12/18 in view of Seed et al. US 5726293, 1998 cited in IDS and Imidazole Elution Buffer, Cold Spring Harbor Protocols. doi:10.1101/pdb.rec356Cold Spring Harb Protoc 2006.
Claim 10-12: Heywood et al disclose a method of purifying an antibody (see figure 14) comprising:
Loading a mixture comprising the antibody to be purified onto an affinity Protein A chromatography resin. See paragraph 9 disclosing elution profile of an antibody from a protein A resin via a pH gradient elution and See paragraph 44 disclosing applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form to a protein A chromatography material wherein said protein A comprises domain D and/or E.
Washing the resin with a wash buffer. See paragraph 70 disclosing wash buffers.
Eluting the antibody with an elution buffer at a pH suitable for releasing the bound antibody from protein A such as pH 3.0 to pH 4.5, pH 3.2 to pH 4.3, pH 3.5 to pH 4 or pH 3.6 or pH 3.9 or pH 3.8. See paragraph 54-55 and 69.
Claim 17: Heywood et al disclose that the method steps a, b and c are performed under dynamic conditions corresponding to add the sample on a protein A column and washing and eluting from the column. See example 1
Claim 18: Heywood et al disclose that the antibody is VH3 domain containing antibody is selected from Fab′, F(ab′).sub.2, scFv, Fab-Fv, Fab-scFv, Fab-(scFv).sub.2, Fab-(Fv).sub.2, diabodies, triabodies, and tetrabodies.
Heywood et al does not disclosing that the elution buffer comprises 0.01M to 1.0M imidazole or an imidazole analogue and a pH of 3 to 5.
Seed discloses that imidazole can be used to disrupt the complex between a protein A and antibody and is a mild elution agent. See under abstract, paragraph 12-14 and 18.
Cold Spring Harbor Protocols (“Cold Spring”) disclose the recipe for 4 elution buffers ranging such as 10 mM, 50 mM, 100 mM and 150 mM imidazole.
It would have been prima facie obvious to a person of ordinary skill in the art as of the effective filing date to have used an elution buffer that comprises 10 mM, 50 mM, 100 mM and 150 mM imidazole and a pH of pH 3.0 to pH 4.5, pH 3.2 to pH 4.3, pH 3.5 to pH 4 or pH 3.6 or pH 3.9 or pH 3.8 to elute the antibody in the method of Heywood et al as taught by Seed, Cold Spring and Heywood, thus resulting in the instant invention with a reasonable expectation of success.
The motivation to do so is that Seed discloses that imidazole can be used to disrupt the complex between a protein A and antibody and imidazole is a mild elution agent and Heywood et al disclose that an elution buffer at a pH suitable for releasing the bound antibody from protein A should be at pH 3.0 to pH 4.5, pH 3.2 to pH 4.3, pH 3.5 to pH 4 or pH 3.6 or pH 3.9 or pH 3.8. Furthermore, Cold Spring Harbor Protocols (“Cold Spring”) disclose the concentration for 4 elution buffers ranging such as 10 mM, 50 mM, 100 mM and 150 mM imidazole.
Claim(s) 14-16  is/are rejected under 35 U.S.C. 103 as being unpatentable over Heywood et al. US 2018/0100007 4/12/18 in view of Seed et al. US 5726293, 1998 cited in IDS and Imidazole Elution Buffer, Cold Spring Harbor Protocols. doi:10.1101/pdb.rec356 Cold Spring Harb Protoc 2006 as applied to claims 10-13 and 17-18 above, further in view of Purifying Challenging Proteins Principles and Methods GE Healthcare (hereinafter referred to as “GE”). Handbook 28-9095-31 AA, 2007.
The combination of Heywood et al and Seed et al and Cold Spring does not disclose that the wash buffer comprises imidazole or 0.01M to 0.5M imidazole and does not disclose that the method further comprises a step of equilibrating the chromatographic resin with an equilibration buffer comprising imidazole prior to loading of the mixture comprising the antibody onto the chromatographic resin.
GE discloses a binding/equilibration buffer comprising 20 to 40 mM imidazole and a wash buffer comprising 20 to 40 mM imidazole. See page 15. The binding buffer can be used to equilibrate an affinity column prior to applying a sample to be purified. See p. 31 teal color paragraph under “purification”.
It would have been prima facie obvious to a person of ordinary skill in the art as of the effective filing date that the method of Heywood et al and Seed et al and Cold Spring as combined can be modified such that the wash buffer can comprise imidazole at a concentration of 20 to 40 mM imidazole and the protein A column can be equilibrated with a binding/equilibration buffer comprising imidazole, thus resulting in the instant invention with a reasonable expectation of success. The motivation to do so is GE health care discloses that the binding buffer serves to equilibrate the column prior to applying the sample and wash buffer washes the column comprising the sample after applying  the sample and prior to elution.
Status of Claims
Claims 10-18 are rejected.

APPLICANT REBUTTAL: Remarks
Claims 10-18 are pending in the subject application. By this Amendment, Applicant has canceled claims 11 and 12 and amended claims 10 and 13. Claim 13 has been amended to attend to an editorial issue It is respectfully submitted that no new matter has been introduced by the amendments made to the claims. Support for the amendments can be found throughout the subject specification and in the claims as originally filed (see, for example, previously pending claims 11-12 and page 5). Entry and consideration of the amendments presented herein is respectfully requested. Accordingly, claims 10 and 13-18 are currently before the Examiner. Favorable consideration of the pending claims is respectfully requested.
Claims 10-13 and 17-18 are rejected under 35 U.S.C. § 103 as obvious over Heywood et al. (U.S. Publication No. 2018/0100007) in view of Seed (U.S. Patent No. 5,726,293) and Imidazole Elution Buffer, Cold Spring Harbor Protocols (2006). Applicant respectfully asserts that the claimed invention is not obvious over the cited references.
Heywood et al. describe a method for purifying an antibody, comprising loading a mixture comprising the antibody to be purified onto a Protein A chromatography resin, washing the resin with a wash buffer, and eluting the antibody with a pH elution buffer at low pH, such as pH 3.0 to pH 4.5. Heywood et al. do not disclose that the elution buffer comprises 0.01 to 1.OM imidazole (or an analogue thereof) and has a pH of 3 to 5. The Office Action argues that the skilled person would have found motivation in Seed to use an elution buffer comprising 10 mM, 50 mM, 100 mM or 150 mM imidazole at a low pH, such as pH 3.0 to pH 4.5, based on the disclosure of Imidazole Elution Buffer/Cold Spring. Applicant respectfully disagrees.
In the claimed method, the affinity chromatography elution buffer has a pH of 3 to 5 and comprises 0.01 to 1.OM of imidazole or an imidazole analogue. The use of such a buffer with a low pH and imidazole (or an analogue thereof) in affinity chromatography results in a reduced elution volume due to improved elution kinetics. This effect is exemplified throughout the as-filed application. For example, faster elution kinetics allowing for a reduction in the number of cycles and elution volume in affinity chromatography has been demonstrated:
- in a MODDE study performed in static binding mode and an IgGi antibody with an elution buffer having a pH of 3.6 or pH 4.0 with 250 mM of imidazole (Example 1, page 14, lines 11 to 15);
- during purification of an IgGI and an IgG4 in dynamic binding mode with an elution buffer having a pH of 3.6 and/or pH 4.0 with 300 mM imidazole (Example 2, page 15, lines 4 to 8 and Figures 2 to 4);
- during purification of a Fab-2x dsscFv in dynamic binding mode with an elution buffer having a pH of 4.0 with 300 mM imidazole (Example 3, page 15, lines 21 to 23 and Figure 5);
- in a MODDE study performed with an affinity chromatography resin in dynamic binding mode and an IgGI antibody with an elution buffer having a pH of 3.5, 3.75 or 4.0 with 250 mM or 500 mM of imidazole (Example 4, pages 15 and 16, lines 10 to 12; and Figure 6); and
- during purification of an IgGI in dynamic binding mode with an elution buffer having a pH of 3.6 with 250 mM dimethyl-1H-pyrazole (Example 5, page 17, lines 10 and 11; Figure 7).
In Seed, antibodies are purified on protein A trisacryl beads and subsequently eluted with solutions comprising 1, 2, 3, 4, or 5M of imidazole, adjusted to a final pH of 6, 7, 8, or 9 (Seed, first example, columns 3 and 4). According to Seed, eluting antibodies with imidazole at a neutral pH is considerably gentler than the more conventional elution techniques, which are based on a change of pH (Seed, column 3, third paragraph). The person of ordinary skill in the art would find no motivation in Seed to add imidazole to the low pH elution buffer used in Heywood et al. to reduce the elution volume in protein A chromatography. In fact, Seed teaches away from the use of a low pH buffer. Based on the disclosure of Seed, the skilled person would use an imidazole-based elution buffer having a neutral pH, i.e. a pH of 6, 7, 8, or 9, in order to avoid denaturation or reduction in activity of the antibody.
Additionally, the best recovery in Seed was observed for those elution buffers which contained 4.0 or 5.OM imidazole (D1, Table 1). It would thus not be obvious to a skilled person to use an elution buffer which comprises the much lower concentration of imidazole, such as a 0.01 to 1.OM imidazole or imidazole-analogue buffer as currently claimed. The deficiencies are not cured by the disclosure of Imidazole Elution Buffer/Cold Spring, since the imidazole elution buffer described therein has a neutral pH of 6.0. Additionally, Imidazole Elution Buffer/Cold Spring does not mention which type of chromatography for which the elution buffer should be used. Accordingly, reconsideration and withdrawal of the rejection under 35 U.S.C. § 103 is respectfully requested.
Claims 14-16 are rejected under 35 U.S.C. § 103 as obvious over Heywood et al. (U.S. Publication No. 2018/0100007) in view of Seed (U.S. Patent No. 5,726,293) and Imidazole Elution Buffer, Cold Spring Harbor Protocols (2006) as applied to claims 10-13 and 17-18 above, and further in view of Purifying Challenging Proteins, GE Healthcare Handbook (2007). Applicant respectfully asserts that the claimed invention is not obvious over the cited references. The defects associated with the combined teachings of Heywood et al., Seed and Imidazole Elution Buffer/Cold Spring have been discussed above. The added teachings of Purifying Challenging Proteins does not remedy these defects in the combined teachings of Heywood et al., Seed, and Imidazole Elution Buffer/Cold Spring. Indeed, the buffer in the Purifying Challenging Proteins document appears to be used for the purification of histidine tagged proteins, not antibodies bound to, for example, Protein A. As such, one skilled in the art would not have looked to that document for buffers suitable for eluting antibodies bound to affinity columns, such as Protein A, Protein G, or Protein L. Accordingly, reconsideration and withdrawal of the rejection under 35 U.S.C. § 103 is respectfully requested.
It should be understood that the amendments presented herein have been made
solely
to expedite prosecution of the subject application to completion and should not be construed as an indication of Applicant's agreement with or acquiescence to the Examiner's position. Applicant expressly reserves the right to pursue the invention(s) disclosed in the subject application, including any subject matter canceled or not pursued during prosecution of the subject application, in a related application.
In view of the foregoing remarks and amendments to the claims, Applicant believes that the currently pending claims are in condition for allowance, and such action is respectfully requested.
The Commissioner is hereby authorized to charge any fees under 37 C.F.R. §§ 1.16 or 1.17 as required by this paper to Deposit Account No. 19-0065.
Applicant invites the Examiner to call the undersigned if clarification is needed on any of this response, or if the Examiner believes a telephonic interview would expedite the prosecution of the subject application to completion.
The undersigned hereby certifies that this correspondence is being transmitted by EFS-Web to the United States Patent and Trademark Office on December 13, 2024.
Respectfully submitted,

NEXT OFFICE ACTION: Notice of Pre-AIA  or AIA  Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Claims 10-18 are pending and are under examination.
The rejection of claim(s) 10-13 and 17-18  under 35 U.S.C. 103 as being unpatentable over Heywood et al. US 2018/0100007 4/12/18 in view of Seed et al. US 5726293, 1998 cited in IDS and Imidazole Elution Buffer, Cold Spring Harbor Protocols. doi:10.1101/pdb.rec356Cold Spring Harb Protoc 2006 is withdrawn. Applicant’s argument are found persuasive.
The rejection of claim(s) 14-16  under 35 U.S.C. 103 as being unpatentable over Heywood et al. US 2018/0100007 4/12/18 in view of Seed et al. US 5726293, 1998 cited in IDS and Imidazole Elution Buffer, Cold Spring Harbor Protocols is withdrawn. Applicant’s argument are found persuasive.
Status of Claims
Claims 10 and 13-18 are allowable.

CITED REFERENCE SUMMARIES: Ref US20180100007A1: Claims:
- 12 . (canceled)
A method for obtaining a human VH3 domain-containing antibody in monomeric form comprising: a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form to a protein A chromatography material wherein said protein A comprises domain D and/or E, under conditions that allow binding of said antibody to protein A, and b) recovering the human VH3 domain-containing antibody in monomeric form,
a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form to a protein A chromatography material wherein said protein A comprises domain D and/or E, under conditions that allow binding of said antibody to protein A, and
b) recovering the human VH3 domain-containing antibody in monomeric form,
wherein the human VH3 domain-containing antibody does not contain an Fc region.
A method for manufacturing a human VH3 domain-containing antibody comprising: a) expressing the antibody in a host cell, b) recovering a mixture containing the antibody, host cells and other contaminants, c) purifying the antibody using at least a protein A chromatography step wherein said protein A comprises domain D and/or E, and d) recovering the human VH3 containing antibody,
a) expressing the antibody in a host cell,
b) recovering a mixture containing the antibody, host cells and other contaminants,
c) purifying the antibody using at least a protein A chromatography step wherein said protein A comprises domain D and/or E, and
d) recovering the human VH3 containing antibody,
wherein the human VH3 domain containing antibody does not contain an Fc region.
A method of separating a human VH3 domain-containing antibody in monomeric form from the antibody in multimeric form comprising: a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form.to a protein A chromatography material wherein said protein A comprises domain D and/or E, b) allowing binding of said antibody to protein A, c) applying an elution buffer that selectively disrupts binding of the antibody in monomeric form, d) recovering the resulting eluate, and optionally e) applying a second elution buffer that disrupts binding of the antibody in multimeric form and recovering this second eluate,
a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form.to a protein A chromatography material wherein said protein A comprises domain D and/or E,
b) allowing binding of said antibody to protein A,
c) applying an elution buffer that selectively disrupts binding of the antibody in monomeric form,
d) recovering the resulting eluate, and optionally
e) applying a second elution buffer that disrupts binding of the antibody in multimeric form and recovering this second eluate,
A method of separating a human VH3 domain-containing antibody in monomeric form from the antibody in multimeric form comprising: a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form to a protein A chromatography material wherein said protein A comprises domain D and/or E, b) allowing binding of the antibody in multimeric form, c) recovering the antibody in monomeric form in the flow-through, and optionally d) applying an elution buffer that selectively disrupts binding of the antibody in multimeric form, and e) recovering the eluate resulting from d);
a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form to a protein A chromatography material wherein said protein A comprises domain D and/or E,
b) allowing binding of the antibody in multimeric form,
c) recovering the antibody in monomeric form in the flow-through, and optionally
d) applying an elution buffer that selectively disrupts binding of the antibody in multimeric form, and
e) recovering the eluate resulting from d);
The method according to claim 13 , wherein a first solution is added to the protein A chromatography material after applying the mixture comprising the human VH3 domain-containing antibody in monomeric and multimeric form such that unbound material is removed in the solution.
The method according to claim 13 , wherein an elution buffer is applied to the protein A chromatography material such that the bound antibody is released.
The method according to claim 13 , wherein the eluate recovered from the protein A chromatography is enriched in monomeric antibody over multimeric antibody with respect to the applied mixture.
The method according to claim 13 , wherein said protein A is native recombinant protein A.
The method according to claim 13 , wherein the VH3 domain containing antibody is selected from Fabâ², F(abâ²) 2 , scFv, Fab-Fv, Fab-scFv, Fab-(scFv) 2 , Fab-(Fv) 2 , Fab-dsFv, diabodies, triabodies, and tetrabodies.
The method according to claim 13 , wherein the VH3 domain-containing antibody comprises at least 2 human VH3 domains
The method according to claim 13 , wherein the VH3 domain-containing antibody specifically binds OX40.
The method according to claim 23 , wherein said VH3 domain-containing antibody comprises: heavy chain CDR1, CDR2 and CDR3 as defined in SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3, respectively; and light chain CDR1, CDR2, and CDR3 as defined in SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6, respectively.
heavy chain CDR1, CDR2 and CDR3 as defined in SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3, respectively; and
light chain CDR1, CDR2, and CDR3 as defined in SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6, respectively.
Ref US20180100007: Claims:
1-12. (canceled)
13. A method for obtaining a human VH3 domain-containing antibody in monomeric form comprising:
a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form to a protein A chromatography material wherein said protein A comprises domain D and/or E, under conditions that allow binding of said antibody to protein A, and b) recovering the human VH3 domain-containing antibody in monomeric form,
wherein the human VH3 domain-containing antibody does not contain an Fc region.
14. A method for manufacturing a human VH3 domain-containing antibody comprising:
a) expressing the antibody in a host cell, b) recovering a mixture containing the antibody, host cells and other contaminants, c) purifying the antibody using at least a protein A chromatography step wherein said protein A comprises domain D and/or E, and d) recovering the human VH3 containing antibody,
wherein the human VH3 domain containing antibody does not contain an Fc region.
15. A method of separating a human VH3 domain-containing antibody in monomeric form from the antibody in multimeric form comprising:
a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form.to a protein A chromatography material wherein said protein A comprises domain D and/or E, b) allowing binding of said antibody to protein A, c) applying an elution buffer that selectively disrupts binding of the antibody in monomeric form, d) recovering the resulting eluate, and optionally e) applying a second elution buffer that disrupts binding of the antibody in multimeric form and recovering this second eluate,
wherein the human VH3 domain-containing antibody does not contain an Fc region.
16. A method of separating a human VH3 domain-containing antibody in monomeric form from the antibody in multimeric form comprising:
a) applying a mixture comprising a human VH3 domain-containing antibody in monomeric and multimeric form to a protein A chromatography material wherein said protein A comprises domain D and/or E, b) allowing binding of the antibody in multimeric form, c) recovering the antibody in monomeric form in the flow-through, and optionally d) applying an elution buffer that selectively disrupts binding of the antibody in multimeric form, and e) recovering the eluate resulting from d);
wherein the human VH3 domain-containing antibody does not contain an Fc region.
17. The method according to claim 13, wherein a first solution is added to the protein A chromatography material after applying the mixture comprising the human VH3 domain-containing antibody in monomeric and multimeric form such that unbound material is removed in the solution.
18. The method according to claim 13, wherein an elution buffer is applied to the protein A chromatography material such that the bound antibody is released.
19. The method according to claim 13, wherein the eluate recovered from the protein A chromatography is enriched in monomeric antibody over multimeric antibody with respect to the applied mixture.
20. The method according to claim 13, wherein said protein A is native recombinant protein A.
21. The method according to claim 13, wherein the VH3 domain containing antibody is selected from Fab′, F(ab′)2, scFv, Fab-Fv, Fab-scFv, Fab-(scFv)2, Fab-(Fv)2, Fab-dsFv, diabodies, triabodies, and tetrabodies.
22. The method according to claim 13, wherein the VH3 domain-containing antibody comprises at least 2 human VH3 domains
23. The method according to claim 13, wherein the VH3 domain-containing antibody specifically binds OX40.
24. The method according to claim 23, wherein said VH3 domain-containing antibody comprises:
heavy chain CDR1, CDR2 and CDR3 as defined in SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3, respectively; and light chain CDR1, CDR2, and CDR3 as defined in SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6, respectively.
Ref US5726293: Claims:
1. A method of isolating an antibody or an antibody fusion protein from a sample, said antibody or said antibody fusion protein comprising a protein A-binding domain, said method comprising:(a) contacting said sample with protein A under conditions which allow affinity complex formation between said antibody or said antibody fusion protein and protein A; (b) separating the complex formed in step (a) from said sample; (c) treating said complex with imidazole in an amount sufficient to release said antibody or said antibody fusion protein from said complex; and (d) recovering said antibody or said antibody fusion protein.
2. The method of claim 1, wherein said antibody is IgG.
3. The method of claim 1, wherein said protein A-binding domain is from an IgG molecule.
4. The method of claim 1, wherein said antibody or said antibody fusion protein comprises a CH2 and a CH3 domain.
5. The method of claim 4, wherein said antibody or said antibody fusion protein further comprises a hinge domain.
6. The method of claim 1, wherein said imidazole is used at a concentration of between 1M and 5M inclusive.
7. The method of claim 6, wherein said imidazole is used at a concentration of 4M.
8. The method of claim 1, wherein said imidazole is used at between pH6 and pH9.