Herein, a facile bottom–up approach for producing nitrogen-doped carbon quantum dots (N-CQDs) was carried out by the hydrothermal treatment of microcrystalline cellulose, in the presence of different nitrogen sources (blank/urea/ammonia water/ethanediamine(EDA)/Hexamethylenetetramine). The result showed that the fluorescence intensity and quantum yields (QYs) of N-CQDs with different nitrogen sources are all higher than that without nitrogen source. Compared with the other three nitrogen sources, N-CQDs prepared by EDA not only have the highest fluorescence intensity but also the largest QYs of 51.39%. Therefore, EDA was chosen as the nitrogen source to prepare N-CQDs. The obtained N-CQDs are uniform spherical particles with a diameter of 2.76 nm. The N-CQDs also exhibit excitation-dependent and long-wave emission properties. The emission range of N-CQDs is 470–540 nm. Moreover, N-CQDs as fluorescent agents successfully acted on purple LEDs (λem = 365 nm) to achieve white LEDs light emission. At the same time, a fluorescent thin layer chromatography plate was successfully prepared using N-CQDs, silica gel G and Sodium carboxymethylcellulose as raw materials. The separation trajectory of mixed sample of Sudan red III and kerosene on the fluorescent TLC plate is obviously clearer than that of the TLC plate.