[WEYDANZ BIRGIT, POMPIATI MARC, SCHAUBMAR ANDREAS, GIESSEL CLAUDIA, GROSSMANN ADELBERT, KRAEMER BRIGITTE, FALKENSTEIN ROBERTO, FUEHRLER NICOLE, HESSE FREDERIKE, GREITHANNER SYBILLE] CH The current invention reports a method for the purification of a not-glycosylated, heterologous polypeptide, which has been recombinantly produced in a prokaryotic cell, wherein the method comprises three chromatography steps of which the first chromatography step selected from i) hydrophobic charge induction chromatography, or ii) hydrophobic interaction chromatography, or iii) affinity chromatography, or iv) ion exchange chromatography, the second chromatography step is selected from i) anion exchange chromatography, or ii) cation exchange chromatography, or iii) hydroxylapatite chromatography, or iv) hydrophobic interaction chromatography, and the a third chromatography step is selected from i) hydrophobic charge induction chromatography, or ii) anion exchange chromatography, or iii) cation exchange chromatography, or iv) hydrophobic interaction chromatography, whereby the first chromatography step is an affinity chromatography in case of polypeptides capable of interacting with metal ligands, the second chromatography step is not a hydroxylapatite chromatography step in case of polypeptides with an isoelectric point below 6.0, and the third chromatography step can be performed in flow-through mode with polypeptides having a low or high isoelectric point.

[HANS STEPHAN, BATHE BRIGITTE, RETH ALEXANDER, CLAES WILFRIED, KRAEMER REINHARD PROF, SEIBOLD GERD, HENRICH ALEXANDER] DE

[FALKENSTEIN, Roberto, WEYDANZ, Birgit, GIESSEL, Claudia, GREITHANNER, Sybille, GROSSMANN, Adelbert, HESSE, Frederike, KRAEMER, Brigitte, POMPIATI, Marc, SCHAUBMAR, Andreas, FUEHRLER, Nicole] Grenzacherstrasse 124 CH-4070 Basel;Wastl-Witt-Strasse 5 80689 München;Neue Heimat 23 82377 Penzberg;Buchener Strasse 25 83646 Bad Tölz;Calvistrasse 13 86971 Peiting;Heimgartenstrasse 1 82436 Eglfing;Ganghoferstrasse 23 80339 München;Woelflstrasse 14a 82377 Penzberg;Waxensteinstrasse 31a 82377 Penzberg;St. Klara Strasse 41 82377 Penzberg;Unterau 44 82444 Schlehdorf The current invention reports a method for the purification of a not-glycosylated, heterologous polypeptide, which has been recombinantly produced in a prokaryotic cell, wherein the method comprises three chromatography steps of which the first chromatography step selected from i) hydrophobic charge induction chromatography, or ii) hydrophobic interaction chromatography, or iii) affinity chromatography, or iv) ion exchange chromatography, the second chromatography step is selected from i) anion exchange chromatography, or ii) cation exchange chromatography, or iii) hydroxylapatite chromatography, or iv) hydrophobic interaction chromatography, and the a third chromatography step is selected from i) hydrophobic charge induction chromatography, or ii) anion exchange chromatography, or iii) cation exchange chromatography, or iv) hydrophobic interaction chromatography, whereby the first chromatography step is an affinity chromatography in case of polypeptides capable of interacting with metal ligands, the second chromatography step is not a hydroxylapatite chromatography step in case of polypeptides with an isoelectric point below 6.0, and the third chromatography step can be performed in flow-through mode with polypeptides having a low or high isoelectric point.

[FALKENSTEIN, Roberto, FLECKS, Birgit, GIESSEL, Claudia, GREITHANNER, Sybille, GROSSMANN, Adelbert, HESSE, Frederike, KRAEMER, Brigitte, POMPIATI, Marc, SCHAUBMAR, Andreas, WEIDLICH, Nicole] Grenzacherstrasse 124, 4070 Basel, CH

[OFTRING ALFRED, KRAEMER KLAUS, NOWAKOWSKY BRIGITTE, OBERMUELLER-JEVIC UTE, TROESCHER ARNULF, STEGMAIER WOLF] DE

[TROETSCHEL CHRISTIAN, KRAEMER REINHARD, BURKOVSKI ANDREAS, BATHE BRIGITTE] DE

[RENOUX BRIGITTE, GESSON JEAN-PIERRE, JACQUESY JEAN-CLAUDE, KRAEMER HANS PETER] FR;FR

[FALKENSTEIN ROBERTO, WEYDANZ BIRGIT, GIESSEL CLAUDIA, GREITHANNER SYBILLE, GROSSMANN ADELBERT, HESSE FRIEDERIKE, KRAEMER BRIGITTE, POMPIATI MARC, SCHAUBMAR ANDREAS, FUEHRLER NICOLE] CH The current invention reports a method for the purification of a not-glycosylated, heterologous polypeptide, which has been recombi-nantly produced in a prokaryotic cell, wherein the method comprises three chromatography steps of which the first chromatography step selected from i) hydrophobic charge induction chromatography, or ii) hydrophobic interaction chromatography, or iii) affinity chromatography, or iv) ion ex-change chromatography, the second chromatography step is selected from i) anion exchange chromatography, or ii) cation exchange chromatogra-phy, or iii) hydroxylapatite chromatography, or iv) hydrophobic interac-tion chromatography, and the a third chromatography step is selected from i) hydrophobic charge induction chromatography, or ii) anion exchange chromatography, or iii) cation exchange chromatography, or iv) hy-drophobic interaction chromatography, whereby the first chromatography step is an affinity chromatography in case of polypeptides capable of in-teracting with metal ligands, the second chromatography step is not a hy-droxylapatite chromatography step in case of polypeptides with an iso-electric point below 6.0, and the third chromatography step can be per-formed in flow-through mode with polypeptides having a low or high iso-electric point.

[ROBERTO FALKENSTEIN, BIRGIT WEYDANZ, CLAUDIA GIESSEL, SYBILLE GREITHANNER, ADELBERT GROSSMANN, FREDERIKE HESSE, BRIGITTE KRAEMER, MARC POMPIATI, ANDREAS SCHAUBMAR, NICOLE FUEHRLER] PROBLEM TO BE SOLVED: To provide a chromatography purification method of a non-glycosylated polypeptide recombinantly produced in a prokaryotic cell.SOLUTION: Provided is a purification of a non-glycosylated, heterologous polypeptide wherein the method comprises three chromatography steps of which the first chromatography step selected from i) hydrophobic charge induction chromatography, or ii) hydrophobic interaction chromatography, or iii) affinity chromatography, or iv) ion exchange chromatography, the second chromatography step selected from i) anion exchange chromatography, or ii) cation exchange chromatography, or iii) hydroxylapatite chromatography, or iv) hydrophobic interaction chromatography, or v) hydrophobic charge induction chromatography, and the third chromatography step selected from i) hydrophobic charge induction chromatography, or ii) anion exchange chromatography, or iii) cation exchange chromatography, or iv) hydrophobic interaction chromatography.

[RENOUX BRIGITTE, GESSON JEAN-PIERRE, JACQUESY JEAN-CLAUDE, KRAEMER HANS PETER] FR;FR