| ELISA Method for Urinary AQP2 Water Channel Protein andits preliminary application in Congestive Heart FailureAbstractObjective:The purpose of this study was to develop a rapid and stable detecting measurement of ELISA for urinary aquaporin-2 (AQP2), to investigate the corelation between urinary AQP2 concentration and hyponatremia in congestive heart failure (CHF).Methods:Rat AQP2 C-terminal peptides(CELHSPQSLPRGSKA) were synthesized and connected with BSA, used to raise polyclonal antibody in rabbit. Parts of the antibodies were attached label with HRP. The direct and sandwich ELISA for urinary AQP2 was tried to developed and applied to CHF rats and patients. The method of ELISA for urinary AQP2 was compared with Western blot analysis.Results:1. The direct ELISA for urinary AQP2 was not sensitive. 2. The sandwich ELISA was able to detect as low as 15.625pmolIml urinary AQP2 with 4.65% and 14.05% of intra-and inter-assay CVs. 3. Urinary AQP2 concentration by this assay was the same regularity as that by Western blot analysis in CHF model rats which were made by ligating the left coronary artery, and higher in large size MI group than in small size MI group. 4. The concentration of urinary AQP2 in CHF patients was higher than health subjects, negative correlative with serum sodium concentration and left ventricular ejection fraction.Conclusion:1. The concentration of urinary AQP2 was too lower to be immobilized by solid phase that the direct ELISA was able to be established. 2. Double antibody sandwich ELISA was established well to detect urine AQP2 concentration in CHF rats and patients. This assay is rapid and simple as compared to Western blot analysis. 3. AQP2 is the target protein in water retention and hyponatremia of CHF. The detection of urinary AQP2 may reflect the severity of water retention and hyponatremia in body. The sandwich ELISA for urinary AQP2 should be a valuable index and a new clinical method to4monitor the water retention and hyponatremia in CHF.
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