Fluorescence lateral flow immunoassays (FL-LFIA) have attracted considerable attention in clinical diagnosis due to their outstanding merits of affordable, sensitive, on-site, and quick detection. However, they are still plagued by significant signal interference, such as autofluorescence and scattered light. The development of high-performance and robust phosphors, i.e., label probes featuring with the character of low/no optical background, remains a great challenge. Herein, we report a novel visualized phosphorescence LFIA (Phos-LFIA), where the composite microspheres, i.e., carbon dots (CDs) covalently embedded in dendritic mesoporous silicon nanoparticles (DMSNs), were designed and selected as the report probes. The obtained CDs@DMSNs revealed uniform morphologies and particle sizes, as well as ultralong (lifetime: 1.14 s, visible for over 8 s to naked eyes) room temperature phosphorescence (RTP) in aqueous solution. As competitive nanotags, CDs@DMSNs were designed for an ultralong phosphorescence-based time-gated LFIA for cardiac troponin I (cTnI) without optical interference. The fabricated Phos-LFIA test strips demonstrated zero-background signal and were applied for highly sensitive cTnI detection in both buffer and a complex serum matrix, with corresponding limits of detection (LODs) of 0.19 and 0.21 ng/mL, respectively. For a clinical validation, the proposed Phos-LFIA revealed an excellent clinical analytical performance (sensitivity: 95.45%, specificity: 88.9%, κ value: 0.85), demonstrating its potential for rapid and accurate diagnosis of myocardial infarction. This work provided a promising background-free probe for FL-LFIA, and it would also open an opportunity for developing highly sensitive screening platforms for other targets through modifying different recognition ligands onto CDs@DMSNs.