The quality of standards for assays is crucial for obtaining consistent results throughout long-term studies and between different studies. Using different batches of these reagents can impact assay validation and reproducibility. Chondrex, Inc.'s ELISA grade collagen undergoes filtration steps to remove polymeric collagen, ensuring a high ratio of monomeric collagen that is ideal for use as standards in sandwich ELISAs and as antigens in indirect ELISAs. This feature is beneficial for establishing reliable ELISA systems. Furthermore, many of Chondrex, Inc.'s kits use ELISA grade collagens as standards and consistently provide reliable and reproducible results for research projects.
The following section explains how the ELISA grade collagen effectively functions in a sandwich ELISA.
In a sandwich ELISA used to detect collagen in samples, typically two antibodies are employed: a capture antibody, which is coated on a solid phase, and a detection antibody, which is conjugated with a probe that can be used to indicate collagen levels. Figure 1 shows an example where five molecules of collagen form one polymeric form (A) and five monomeric forms (B). These forms are used as standard collagen in the sandwich ELISA. The capture antibodies bind to the polymeric collagen in samples, but only two detection antibodies can bind to their epitopes on the collagen because the epitopes inside the polymeric collagen are not exposed. As a result, the assay displays two signals. Conversely, the capture antibodies can bind all five monomeric collagens, allowing five detection antibodies to bind to each collagen molecule respectively. This results in the assay displaying five signals. Variations in the composition of monomeric and polymeric collagen in the batches used for standards can cause fluctuations in the assay results, particularly in the standard curve, significantly affecting the calculated collagen levels in samples.
To obtain absolute values of collagen levels in samples, monomeric collagen (ELISA grade collagen) must be used as a standard in ELISA.
