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Methodology of generation and purification of anti-beta 2 glycoprotein I antibodies

Marcelo Alarcon1, Eduardo Fuentes1, Ximena Maldonado2

  • 1Thrombosis Research Center, Medical Technology School, Department of Clinical Biochemistry and Immunohaematology, Faculty of Health Sciences, Research Center for Aging, Universidad de Talca, Talca, Chile.

Methodsx
|May 14, 2019
PubMed

In this Method Article we are showing the methodology for generation and purification of Anti-Beta 2 Glycoprotein I (β2GPI) antibodies. First β2GPI was purified from human plasma, and recognized by Western Blot and anti-β2GPI antibodies of serum from patients with antiphospholipid syndrome (APS). The C57BL/6 mice were immunized intraperitonealy with 150 μg of protein in adjuvant (β2GPI or bovine serum albumin) on days 1, 8 and 14. Then the anti-β2GPI antibodies were purified by affinity chromatography (Affi-Gel protein A sepharose) and affinity column using human β2GPI coupled to CNBr-activated Sepharose 4B. Titles of anti-β2GPI antibodies were determined by ELISA assays. •We purified β2GPI with great efficacy and that is recognized antigenically by serum from patients with SAP or an anti-β2gpi antibody.•We found that our purified antibody had 13 fold increased activity in ELISA test compared with the control and in Western Blot recognized with β2GPI (reference and purified).

Keywords:
AntibodiesAntiphospholipid syndromeBeta 2 glycoprotein IGeneration and purification of Anti-Beta 2 Glycoprotein I antibodiesPurification

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