The quantitative analysis by matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) is a challenge due to the poor reproducibility originating from the heterogeneity of the matrix-analyte crystals. Polystyrene (PS) colloidal spheres have superior monodispersed property and can self-assemble to form photonic crystals. With the assistance of PS spheres, a uniform matrix-analyte cocrystal was constructed for the quantitative analysis of plasma lysophosphatidylcholines (LPCs). The matrix and the solvent in MALDI MS analysis were optimized, and the reproducibility and the accuracy were investigated in detail. This is the first report about the very simple method of PS spheres-assisted MALDI MS for quantitative analysis, where it is believed that this approach will greatly expand the applications of MALDI MS.