Beta-cyfluthrin, as a synthetic pyrethroid, has been widely used in cigarette manufacturing factories in China to control Lasioderma serricorne (F.) (Coleoptera: Anobiidae). In this study, spray toxicity bioassays and filter paper residual contact toxicity bioassays were conducted to investigate the beta-cyfluthrin sensitivity level of five field strains of L. serricorne collected from cigarette manufacturing factories in China. Bioassay results indicated that five field strains had developed different levels of resistance to beta-cyfluthrin with RR₅₀ of 3.51–10.20 at 2 hr after application and 4.05–49.50 at 24 hr after application in spray toxicity bioassays, and RR₅₀ of 4.74–14.47 at 2 hr exposure in filter paper residual contact bioassays. In addition, we examined CarE, GST, and CYP450 enzyme activity and content of L. serricorne adults and larvae. Enzyme-linked immunosorbent assay results suggested that there was no significant difference in GST, CYP450, and CarE content of L. serricorne adults between field strains and reference sensitive strain. Biochemical assay results indicated that CYP450 activity of L. serricorne adults and larvae of five field strains was significantly higher than that of reference sensitive strain, with increased CYP450 activity of 1.08–1.82-fold in adults and 1.08–2.12-fold in larvae. The results implied that elevated CYP450 activity may contribute to metabolic resistance of L. serricorne to pyrethroid. Our study indicated that there was no clear evidence that the enhanced CarE and GST activity was associated with pyrethroid resistance of L. serricorne.