Immunoassay of Modified Forms of Human Low Density Lipoprotein in Isolated Circulating Immune Complexes

Modified lipoproteins are able to induce inflammatory reactions through innate immunity pathways and are immunogenic, leading to an autoimmune response that results in the formation of proinflammatory immune complexes. The measurement of circulating oxidized lipoproteins and corresponding antibodies has, therefore, been proposed as an approach to assess the risk for complications in patients with diabetes and for the risk of cardiovascular disease in the general population. However, the majority of modified low density lipoprotein (LDL) in the peripheral circulation exists in the form of immune complexes, and this is a significant obstacle for the measurement of modified LDL and the corresponding antibodies. In this manuscript, we describe in detail the methodology developed by our group for isolation and fractionation of circulating immune complexes (IC), allowing the accurate assay of different LDL modifications. This approach has resulted in several studies showing that the levels of modified LDL are risk factors with a stronger association to diabetic retinopathy, nephropathy, and macrovascular disease. Ongoing research is focused on evaluating the predictive power of modified LDL levels for the development or progression of atherosclerotic cardiovascular disease in other patient populations and on the simplification of the assay to make it more applicable to diagnostic laboratories.