Sensitive and Selective Immunofluorescence Assay for CA15-3 Detection Using Fluorescein Derivative A10254
Background: Carbohydrate antigen 15-3 (CA15-3), a specific breast cancer-related biomarker in serum, can provide direct information to monitor a patient's postoperative status and can predict recurrence and metastasis of breast cancer. The conventional Enzyme-Linked Immunosorbent Assay (ELISA) is an effective approach for CA15-3 detection, but there is a high limit of detection which is more than 1 U mL-1. Therefore, a convenient, highly sensitive and specific CA15-3 detection method is needed for earlier prognoses of breast cancer. Objective: In this work, we aim to develop a novel sensitive and selective immunofluorescence assay for the highly effective detection of CA15-3. Method: We pre-coat the antibody I onto the solid phase to capture antigen CA15-3. The A10254- labeled antibody II identified by electrophoresis and fluorescence spectra recognized the antigen CA15-3 and formed the sandwich format. The fluorescence signal of A10254 directly relating to the amount of antigen could be detected and quantified by the microplate reader excited at 440nm. Results: The obtained method for CA15-3 detection showed lower limits of detection (0.11 and 0.18 U mL-1) in a linear range of 0.25-14 U mL-1 in the buffer and 2% serum, respectively. This assay exhibited a good specificity and reproducibility in 2% serum with lower relative standard deviations in different tested concentrations. Conclusion: The developed immunofluorescence assay enhanced the performance of the conventional CA15-3 ELISA kit. Moreover, the acceptable accuracy and good reproducibility in diluted serum indicated the assay has great potential for the diagnosis of breast cancer in clinical practice.
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