As a promising analytical technique in recent years, surface-enhanced Raman spectroscopy (SERS) has received extensive attention due to its low limit of detection, high sensitivity and high specificity. Despite its tremendous potential, SERS was not widely applied in quantitative analysis of chemical and biological samples in the past years. However, the explosive development of nanotechnology and nano-fabrication has assisted the development of SERS as a quantitative analysis tool. As the enhancement of Raman scattering strongly depends on nanoscale surface morphology of the enhancing surface and can be easily influenced by other factors in an experiment, it is still a challenge to obtain reliable results comparable to those obtained from state-of-the-art analysis methods. The fabrication of three kinds of enhancing media including traditional solid substrates, colloidal nanoparticles and plasmonic nanostructures based on nano-fabrication and their respective advantages and drawbacks for quantitative SERS detection are summerized in this review. Furthermore, how to improve the sensitivity and reliability is investigated in aspects of molecular orientation, excitation wavelength, internal standard and data analysis. Meanwhile, several successful cases of quantitative SERS detection are presented. Finally, applications and prospects of its future researches are proposed.