Systems, materials, and methods for reversed-phase high performance liquid chromatography (rp-hplc) identification of multi-specific molecules
AssigneeJanssen Biotech, Inc.
InventorsJames Martosella, Jessika S. Feliciano-Cardona, Gulnur Elove
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Abstract
Provided herein are high-pressure liquid chromatography (HPLC) methods for detecting multi-specific molecule formation after a multi-specific molecule manufacturing or development process. The HPLC methods provide fast, quantitative, real-time processing of multi-specific molecule formation.
Claims29
A method of detecting formation of a multi-specific molecule, the method comprising:
- a. obtaining a multi-specific molecule sample;
- b. obtaining a reversed-phase high performance liquid chromatography (RP-HPLC) column;
- c. contacting the multi-specific molecule sample in a polar aqueous mobile phase A with the RP-HPLC column, wherein the polar aqueous mobile phase comprises an ion-pairing agent;
- d. contacting an organic non-polar mobile phase B with the RP-HPLC column, wherein the organic non-polar phase comprises an ion-pairing agent;
- e. eluting the multi-specific molecule sample;
- and
- f. monitoring the amount of multi-specific molecule formation in the eluted multi-specific molecule sample.
A method of detecting formation of a multi-specific antibody while performing a manufacturing process to produce a multi-specific antibody, the method comprising:
- a. performing a manufacturing process to produce a multi-specific antibody, wherein the manufacturing process results in a multi-specific antibody sample comprising an amount of a multi-specific antibody;
- b. setting up a reversed-phase high performance liquid chromatography (RP-HPLC) column;
- c. injecting the multi-specific antibody sample in a polar aqueous mobile phase A in the RP-HPLC column, wherein the polar aqueous mobile phase comprises an ion-pairing agent;
- d. applying an organic non-polar mobile phase B to the RP-HPLC column to form a gradient elution, wherein the organic non-polar mobile phase comprises an ion-pairing agent;
- e. eluting the multi-specific antibody sample;
- and
- f. monitoring the amount of multi-specific antibody formation in the eluted sample.
A method of detecting formation of a bispecific antibody while performing a controlled FAB arm exchange (cFAE) to produce a bispecific antibody, the method comprising:
- a. performing a controlled FAB arm exchange (cFAE) to produce a cFAE sample, wherein the cFAE sample comprises an amount of a bispecific antibody;
- b. setting up a reversed-phase high performance liquid chromatography (RP-HPLC) column;
- c. injecting the cFAE sample in a polar aqueous mobile phase A in the RP-HPLC column, wherein the polar aqueous mobile phase comprises an ion-pairing agent;
- d. applying an organic non-polar mobile phase B to the RP-HPLC column to form a gradient elution, wherein the organic non-polar mobile phase comprises an ion-pairing agent;
- e. eluting the cFAE sample;
- and
- f. monitoring the amount of bispecific antibody formation in the eluted sample.
A system comprising a reversed-phase high performance liquid chromatography (RP-HPLC) member for detecting formation of a multi-specific molecule, the system comprising:
- a. a multi-specific molecule sample in a polar aqueous mobile phase A, wherein the polar aqueous mobile phase comprises an ion pairing agent;
- b. a RP-HPLC column;
- c. an organic non-polar mobile phase B, wherein the organic non-polar mobile phase comprises an ion pairing agent;
- wherein the RP-HPLC column is configured to be contacted with the multi-specific molecule sample in polar aqueous mobile phase A, and
- wherein the RP-HPLC is further configured to be contacted with the organic non-polar mobile phase B to elute a multi-specific molecule sample, wherein an amount of multi-specific molecule formation can be identified in the eluted multi-specific molecule sample.
A means for reversed-phase high performance liquid chromatography (RP-HPLC) detection of multi-specific molecule formation comprising:
- a. a multi-specific molecule sample in a polar aqueous mobile phase A, wherein the polar aqueous mobile phase comprises an ion pairing agent;
- b. a RP-HPLC column;
- c. an organic non-polar mobile phase B, wherein the organic non-polar mobile phase comprises an ion-pairing agent;
- wherein the RP-HPLC column is configured to be contacted with the multi-specific molecule sample in polar aqueous mobile phase A, and
- wherein the RP-HPLC column is further configured to be contacted with the organic non-polar mobile phase B to elute a multi-specific molecule sample, wherein an amount of multi-specific molecule formation can be identified in the eluted multi-specific molecule sample.
Dependencies are inferred from the claim text.