Background: Aldo-ketoreductase family 1 member B10 (AKR1B10) is a novel prognostic predictor and therapeutic target for colorectal cancer (CRC), and enzyme-linked immunosorbent assays (ELISAs) and electrochemiluminescence (ELC) assays are sample-consuming and high-cost methods. Therefore, it is very necessary to develop a new, simple, and fast yet highly sensitive and specific method for the detection of AKR1B10 in serum. Semiconducting quantum dots (QDs) possess a high fluorescence quantum yield, stability against photobleaching, and size-controlled luminescence properties; thus, they are suitable for photoelectrochemical tumor marker detection, especially in complex biological samples. However, CdTe/CdS QDs have not been applied for the detection of AKR1B10 in serum.
Methods: AKR1B10 in peripheral blood has been established using anti-AKR1B10-conjugated CdTe/CdS QDs and measurements. The assay sensitivity was determined by measuring the quenched fluorescence intensity of AKR1B10 at 0.5, 1, 2, 5, or 10 ng/mL in phosphate-buffered solution (PBS) or 0.25%, 0.5%, 1.0%, 2.0%, or 5% human serum diluted in PBS. The assay was optimized under different pH values (7.00 - 7.40) for different reaction durations (10 - 60 minutes). The specificity of anti-AKR1B10-QDs was determined by testing the inhibition of AKR1B10 activity with carcinoembryonic antigen (CEA), immunoglobulin G (IgG), or alpha-fetoprotein (AFP), each at 1 ng/mL.
Results: Under the optimized incubation time (30 minutes) at room temperature and optimal pH (7.1 - 7.2), a correlation between the decreased fluorescence intensity of anti-AKR1B10-conjugated CdTe/CdS QDs and the concentration of AKR1B10 in the range from 0.05 to 100 ng/mL was established. The assay was sensitive for the detection of AKR1B10 in the range from 0.05 to 100 ng/mL, and the detection limit was 0.02 ng/mL. The assay presented a high specificity because the anti-AKR1B10-conjugated CdTe/CdS QDs only reacted with AKR1B10 in the sera in the presence of CEA, IgG, or AFP.
Conclusions: In conclusion, the immunofluorescence assay to detect AKR1B10 in serum using anti-AKR1B10-conjugated CdTe/CdS QDs was simple and fast yet presented high sensitivity and specificity. Our findings provide a promising tool for the early prediction of CRC.