(m, r Nephrin ELISA, Strip Plate) The Human Nephrin kit contains all of the components...
Efficiently assess glomerular integrity in studies of diabetes, renal disease, and other pathogenic processes with our ready-to-use Nephrin ELISA kits.
FOR RESEARCH USE ONLY
Used as a biomarker for kidney injury and dysfunction, urinary nephrin levels are an important research tool for studying renal diseases and diabetes. Advance these studies with our Nephrin ELISA Kits, optimized for nephrin detection in human and rat urine samples. Part of our reliable Exocell ELISA portfolio, both human and rat kits are configured as competitive ELISAs for specific and robust detection without the need for extensive sample pre-processing. Detection is using an indirect method, with a primary antibody raised in mouse against the N-terminal portion of the nephrin protein from either rats (for the rat ELISA) or humans (for the human ELISA), and an anti-mouse secondary antibody conjugated to horseradish peroxidase (HRP). Measurement is accomplished via a colorimetric substrate that absorbs at 450 nm (note that because these are competitive assays, increasing nephrin levels result in decreasing signal).
The Nephrin ELISA Kits are part of our Exocell portfolio and deliver the same reliable results and expert support that Exocell customers know to expect, including reagents that are ready-to-use either as is or with only moderate dilution. Quantitation is simplified through the use of the Creatinine Companion, which enables normalization to creatinine levels to account for variable urine concentration.
What’s a competitive ELISA?
As the name suggests, in a competitive ELISA, the amount of analyte in a sample is determined via a competitive reaction where increasing amounts of analyte result in decreased signal. Here’s how it works:
In a competitive ELISA, the wells of the plate are pre-coated with set amounts of purified analyte which remains immobilized in the well. In the absence of sample, addition of the detection antibody results in maximum signal. However, when sample is added, any analyte present will compete with the immobilized analyte for binding to the detection antibody, resulting in decreased antibody remaining the wells after washing and, thus, decreased signal in the presence of analyte.
Why use a competitive ELISA?
Competitive ELISAs deliver highly quantitative results yet are relatively quick to complete as they require little to no sample processing. Because detection is via a competitive reaction, the impact of low affinity, off-target binding is minimized enabling the use of fairly crude samples and rendering the assay less sensitive to sample dilution and sample matrix effects. Competitive ELISAs are also more reproducible, with low variability between duplicates and from assay-to-assay, and they have the flexibility to work with small antigens, antigens with few epitopes, and unpurified or polyclonal antibodies.
What is direct vs. indirect detection?
Direct versus indirect detection refers to how HRP is introduced into the assay. With direct detection, HRP is conjugated directly to the antibody that recognizes the analyte. This is the simpler and quicker of the two methods as only a single antibody is needed. With indirect detection, a primary antibody recognizes the analyte, and then a secondary antibody conjugated to HRP recognizes the primary antibody. The addition of the secondary antibody adds wash steps, increasing the time and labor involved in the assay workflow as well as adding steps where variability can be introduced.
Nephrin is a transmembrane protein expressed in renal glomerular podocytes, the visceral epithelial cells that line the outer aspect of the glomerular basement membrane, and is an integral component of the slit diaphragm which connects adjacent podocyte foot processes. The slit diaphragm regulates the passage of plasma proteins across the glomerular filtration barrier and is critical for kidney function. Mutation of the nephrin gene, altered nephrin production, and abnormal podocyte function lead to proteinuria in diseases affecting the glomerulus such as nephrotic syndrome and diabetes.
Podocyte injury may be accompanied by shedding of the nephrin protein and/or of podocytes into the urine, and studies in experimental animals and in human diabetes support the hypothesis that nephrinuria is a marker for, and may reflect severity of, glomerular filtration dysfunction. ELISA measurement of urinary nephrin may provide an avenue for detection of early renal dysfunction and/or for assessing response to therapeutic interventions in experimental animal and clinical research.