In this study, the effect of cell-free aulture supernatant of Lactobacillus plantarum-Y42 on the biofilm formation of Listeria monocytogenes (LM) was assessed. Minimum inhibitory concentration (MIC) of L. plantarum-Y42 cell-free culture supernatant against planktonic LM was determined using the Resazurin staining method. Then the effect of sub-MICs of L. plantarum-Y42 cell-free culture supernatant on the swarming motility and biofilm formation of LM were tested. MIC of L. plantarum-Y42 cell-free culture supernatant against LM was 60mg/mL. When LM was exposed to L. plantarum-Y42 cell-free culture supernatant at the sub-MICs of 0,1/32,1/16,1/8,1/4,1/2MIC, respectively, the corresponding swarming diameters of LM were 81.15,8.31,5.35,9.51,4.91, and 33.09mm, respectively. The inhibition rates of biofilm formation were 0,2.90%, 30.81%, 40.75%, 39.93%, and 12.15%, respectively. Sub-MICs of L. plantarum-Y42 supernatant did not inhibit the normal growth and proliferation of LM, but significantly inhibited the biofilm formation of LM.