The invention provides a single particle layer nano-diamond film and a preparation method thereof. The preparation method comprises the following steps: polishing a monocrystalline silicon piece by using a diamond, and then cleaning and drying to obtain a monocrystalline silicon substrate; putting the monocrystalline silicon substrate in hot filament chemical vapor deposition equipment; by taking acetone as a carbon source, bringing acetone to a reaction chamber in a hydrogen bubbling manner, wherein the flow rate of hydrogen to acetone is 200: 90, the distance from a hot filament to the monocrystalline silicon substrate is 7mm, the reaction power is 1600-2300W, and the working air pressure is 1.63Kpa, no bias voltage is applied in the reaction process, the film growing time is 10-50 minutes; and after growth, dropping the temperature and cooling under a condition of not introducing hydrogen, thus obtaining the single particle layer nano-diamond film which is 300-700nm thick. The film is relatively strong in Si-V light-emitting property and has a broad application prospect in the field of single-photon sources, quantum information processing, optoelectronic devices, electrochemical electrodes, biomarkers, semiconductor apparatuses, field emission displays and the like.