Comparison of Two Strategies of Analysis of Urinary Protein Composition for the Diagnosis and Follow-Up of Renal Diseases

Abstract

Proteinuria analysis is necessary to detect the early stages of kidney disease before the estimated glomerular filtration rate deteriorates and to monitor the progression of treated kidney disease. Electrophoresis is often the first orientation test, although this test is time-consuming and its interpretation may be subjective. Two types of electrophoresis gel for urinary proteins are available: (1) high-resolution (HR) agarose gel and (2) agarose gel combined with immunological detection of specific urinary proteins after their electrophoretic migration (UP). As the former is known to provide the best results for the quantification of monoclonal protein and the latter for its characterization, we only investigated methods for determining the type of kidney damage in our study. Therefore, the aim of our study was to compare two strategies for proteinuria typing: UP gel to HR gel with quantification of specific proteins (albumin, eventually transferrin, α1-microglobulin, eventually β2-microglobulin, immunoglobulin G (IgG), and α2-macroglobulin if blood is present), and HR gel allowing the visualization of RBP, β2-microglobulin, and transferrin. The two methods were comparable in detecting abnormal excretion of tubular markers (α1-microglobulin and β2-microglobulin), nonselective glomerular markers such as IgG, and post-renal marker (α2-macroglobulin). The results differed for albumin, whose limit of detection was 25 times lower than the limit of quantification by immunoassay, leading to false positives and no differentiation between low and high excretion of albuminuria. In conclusion on UP gel, when proteinuria typing is prescribed without investigating monoclonal protein, we recommend carrying out immunoassays for specific proteins (e.g., albumin, α1-microglobulin, eventually β2-microglobulin, and IgG). In a context of associated monoclonal protein investigation, tubular and glomerular damage markers (excluding albumin) can be interpreted on a UP gel. If blood is present, α2-macroglobulin must be measured using immunoassays to determine the post-renal origin of proteinuria using the ratio α2-macroglobulin/albumin.

Keywords: albuminuria; chronic kidney disease; electrophoresis; proteinuria typing; quantitative immunoassays.

FIGURE 1

Example of UP gel electrophoresis. The first lane “ELP” corresponds to all migrated urinary proteins, which were revealed by only staining. The revealed proteins on the other lanes correspond to colored antigen–antibodies complex: Tub, tubular proteinuria revealed with using α1‐microglobulin, RBP, and β2‐microglobulin antibodies; Alb/A2M, albumin/α2‐macroglobulin; GAM, IgG, IgA, IgM; DE, IgD, IgE; Kl/f, bound/free kappa; Ll/f, bound/free lambda.

FIGURE 2

Proposed flowchart for interpreting albumin revealed by antibodies on UP gels, applied to our study. A2M, α2‐macroglobulin; Alb, albumin; TRF, transferrin.

FIGURE 3

Suggested flowchart for proteinuria typing depending on the laboratory's possibilities or choice of immunoassay kits for specific proteins. There are three possible scenarios, each of which includes albumin ± α2‐macroglobulin obtained by immunoassay. (Left) Without access to electrophoresis, the laboratory must perform A1M and IgG assays to be able to type proteinuria. However, this approach will not detect monoclonal protein. (Middle) UP gels provide additional information about specific proteins in order to type the proteinuria and detect the monoclonal component. (Right) All specific proteins must be analyzed using immunoassays and HR gels, providing additional information about other specific proteins and quantifying the monoclonal component. A1M, α1‐microglobulin; A2M, α2‐macroglobulin; Alb, albumin; IA, immunoassay; IgG, immunoglobulin.