A nanocomposite of multi-walled carbon nanotubes/gold nanoparticles/poly-melamine (MWCNTs/AuNPs/PM) was designed using layer-by-layer self-assembled method on glassy carbon electrode (GCE) by electrochemical deposition to construct an electrochemical sensor for sensitive detection of nitrite. First, a layer of MWCNTs was modified on electrode, and then gold nanoparticles and melamine were in-situ polymerized onto MWCNTs through self-assembled technique to form GCE/MWCNTs/AuNPs/PM. MWCNTs have large specific surface area, which increased the number of gold nanoparticles deposited on MWCNTs. Meanwhile, the doping of gold nanoparticles also improved the polymerization of melamine. The synergistic interaction of nanocomposite further improved the catalytic effect on nitrite. Under optimized conditions, the detection range for nitrite was from 0.4 to 1475 μM and the detection limit was 0.041 μM. Through the detection of nitrite in food samples, the recovery rates were from 93.16% to 108.68%. Therefore, the method can be used as a practical platform for nitrite detection in food.