o:35964 Simultaneous Isolation and Purification of Transferrin and Immunoglobulin G from Human Serum-A New Biotech Solution en ABSTRACT A fast and simple biotech method is presented for the simultaneous isolation and purification of transferrin (Tf) and immunoglobulin G (IgG) from the same pool-sample of human serum, yielding >98% pure proteins. Serum sample preparation was achieved by precipitation with ethacridine lactate (rivanol). Protein purification was performed with AKTA Avant 150 FPLC, using a Resource Q column. Three different buffers at pH 6.2 (MES, phosphate, and Bis-Tris) were tested. Isolated and purified proteins retained their native 3D structure, as shown by spectrofluorimetric measurements. Tf functionality was preserved, as confirmed by the retention of both the iron binding capacity and its ability to interact with the transferrin receptor (immunofluorescent staining), as well as the immunogenicity of IgG, as shown by Western blot analysis with immunodetection. The formation of IgG aggregates was avoided. This biotech method is a rapid, simple, and time-saving alternative to other methods for the isolation of extremely pure IgG and Tf, while it is also the only method so far described for their simultaneous isolation. biotherapeutics; isolation and purification; ion-exchange; IgG; transferrin 1552099 10.3390/molecules30050993 2025-02-24T20:28:55.734Z 44 yes 46 Danilo Četić University of Belgrade, Institute for the Application of Nuclear Energy INEP, Belgrade, Serbia 0000-0001-9705-3237 Goran Miljuš University of Belgrade, Institute for the Application of Nuclear Energy INEP, Belgrade, Serbia doktor biohemijskih nauka person 0000-0001-6417-8711 Zorana Dobrijević University of Belgrade, Institute for the Application of Nuclear Energy INEP, Belgrade, Serbia doktor bioloških nauka person 0000-0002-4652-7719 Nikola Gligorijević University of Belgrade, Institute of Chemistry, Technology and Metallurgy, Belgrade, Serbia person Aleksandra Vilotić University of Belgrade, Institute for the Application of Nuclear Energy INEP, Belgrade, Serbia doktor bioloških nauka person 0000-0002-0898-5401 Olgica Nedić University of Belgrade, Institute for the Application of Nuclear Energy INEP, Belgrade, Serbia doktor biohemijskih nauka person 0000-0003-2042-0056 Ana Penezić University of Belgrade, Institute for the Application of Nuclear Energy INEP, Belgrade, Serbia doktor biohemijskih nauka person 0000-0002-2539-4507 application/pdf 3624315 https://phaidrabg.bg.ac.rs/o:35964 no yes 16 70 92000001 11A29 11A2905 11A24 molecules 30 5 MDPI, Basel, Switzerland 2025