Lateral flow assay is a new in vitro diagnostic technique developed on the basis of monoclonal antibody technology, colloidal gold immunochromatography and new material technology in the 1990s, with rapid, simple, single detection, low-cost features. It is now widely used in medical test, food safety, environmental, agricultural and veterinary safety, inspection and quarantine, forensic fields, etc.
Figue1. Rapid immunochromatographic test strip
The specific antigen or antibody was immobilized on the NC membrane (nitrocellulose membrane) with a large pore size microporous membrane as a carrier. When the sample was added to the sample at one end of the test strip, on the adsorbent sample pad, it had specific immune response with the binding of colloidal gold or microspheres labeled reagent via lateral movement by capillary action; then moved to the NC membrane, and then captured by antigen or antibody fixed in the NC membrane surface, and gathered in the detection area, then the density of the light-reflecting signal was visually obtained by visualizing the nitrocellulose surface marker (colloidal gold or latex particles). Other unbounded markers crossed the test strip and flow into the absorbent pad to achieve the purpose of automatic separation.
With the rapid development of immunoassay technology, the quantity, high sensitivity, multiple detection and system integration have become a new research hotspot in the field of IVD. Compared with the traditional ELISA and qualitative lateral colloidal gold chromatographic determination, the quantitative lateral chromatographic determination based on microspheres has many advantages such as good stability, wide linear range and high sensitivity. The commonly used materials for preparing Estapor nanospheres are polystyrene (PS) or composites composed of polystyrene and other compounds such as divinylbenzene.
For microspheres with no functional groups on the surface, the antigen or antibody is immobilized on the surface of the microspheres by passive adsorption, and the process is simple. For microspheres modified with functional groups, the antigen / antibody can be immobilized on the surface of the microspheres by covalent cross-linking. There are two antigen/antibody covalent cross-linking methods, one is two-step, in which cross-linked microspheres need to activate, for example, -COOH carboxyl modified microspheres can be activated with NHS / EDC; the other is a one-step, in which microspheres cross-linked only need to change the pH value of the buffer to get activated; the operation is very convenient, and the chloromethyl, toluenesulfonyl, epoxy-modified microspheres also belong to this category. Antigen or antibody conjugated to the surface of the microspheres can be removed by centrifugation, dialysis or ultrafiltration to remove the uncrosslinked antibody / antigen and then re-dispersed using ultrasound.