The concept of green chemistry was advocated, and near-infrared (NIR) red fluorescence emission carbon dots (RCdots) were prepared using the hydrothermal method with blood orange. RCdots adsorbed alpha-fetoprotein aptamers (AFP-aptamers) onto the surface through π-π stacking, and a photoinduced electron transfer (PET) effect occurred, leading to the fluorescence quenching of RCdots. However, AFP has a stronger affinity with AFP-aptamer, causing the AFP-aptamer to be removed from the surface of RCdots, resulting in the restoration of RCdots' fluorescence. Based on this principle, an aptamer-based NIR fluorescence probe was developed to specifically and sensitively detect AFP, and it was applied to the sensitive quantitative detection of AFP in human serum. The experimental results indicated that the linear range of AFP detection was 0.01 ng/mL ~ 105 ng/mL, and the detection limit was 3.5 pg/mL. To further verify the feasibility of this method, the aptamer-based NIR fluorescence probe was also used in the imaging of AFP in hepatoma cells (HepG2 group), normal hepatocyte group (HL-7702 group), and other tumor cell groups in the experiment. Additionally, the AFP in mouse hepatoma tumor cells was measured analyzed by fluorescence imaging in vivo. The results revealed that the fluorescence intensity of the probe in HepG2 tumor cells with high AFP content was significantly higher than that in normal cells and other tumor cell groups. Furthermore, the fluorescence intensity emitted by the probe at the mouse HepG2 tumor site was significantly higher than that in normal tissue, and the fluorescence intensity enhanced with the increase of AFP concentration. These results indicate that the aptamer-based NIR fluorescence probe provides valuable technical reference for developing green and sustainable production of characteristic agricultural sources. Meanwhile, it can be used for the diagnosis of early liver cancer through AFP detection and imaging. It can also serve as an ideal aptamer-based NIR fluorescence probe in the study of related tumor treatment under the guidance of in vivo imaging, and offers a promising application prospect in the fields of clinical diagnosis, biological analysis, and life science research.