Point of care testing (POCT) of cholesterol in blood serum via a moving reaction boundary electrophoresis titration chip

Abstract

Cholesterol (CHO) in human blood is one of the most frequently and crucially quantified substances in diagnostic laboratories. However, visual and portable point of care testing (POCT) methods have been rarely developed for the bioassay of CHO in blood samples. Here, we developed an electrophoresis titration (ET) model, a chip device of ∼60 grams, and a quantification method for the POCT of CHO in blood serum based on a moving reaction boundary (MRB). In this model, the selective enzymatic reaction is integrated with an ET chip for visual and portable quantification. At first, CHO reacted with cholesterol oxidase (CHOx) in the anode well, producing H₂O₂ and cholest-4-en-3-one in the solution. H₂O₂ further oxidized the colorless and chargeless leucocrystal violet (LCV) dye into violet colored positively charged crystal violet (CV⁺) and, under the influence of the electric field, the CV⁺ migrates in the ET channels and is titrated by the alkali of sodium hydroxide immobilized in the ET channels. The length covered by the MRB was measured as a function of the CHO content. The relevant experiments validated the feasibility of the model and method. Furthermore, the experiments revealed the high selectivity, portability, and visuality of the ET-MRB model, device, and method. Finally, the experiments showed a fair sensitivity of LOD of 5 μM, good linearity of 10–1000 μM (r² = 0.9919), fair stability (intra-day RSD of less than 5.09% and an inter-day RSD of less than 6.36%), and high recovery (99.4–105%). All the data and results indicate the potential of the ET-MRB model, chip device, and method for POCT of CHO in human blood samples.