Lyme Disease Antibody Generation Protocol

Swiss-Webster mice were purchased from Taconic (Hudson, NY). Ixodes scapularis tick larvae were obtained from BEI Resources (Manassas, VA). B. burgdorferi-infected nymphs were generated as described in Generation of Nymphal Ticks Carrying B. burgdorferi. All B. burgdorferi, B. afzelii, and B. garinii strains used in this study were grown in BSK-II complete medium (Table S1). To generate the antisera against different Lyme borreliae species, B. burgdorferi strain B31-A3, B. afzelii strain VS461-JL, or B. garinii strain ZQ1 was intra-dermally introduced into mice (10⁶ bacteria per mouse). At 21 days post-infection, an ear biopsy (one biopsy per mouse) was collected to determine the infectivity of the particular strain used to infect that mouse using quantitative PCR (qPCR; see Quantitative PCR and the previous description^{53 (link)}). Sera were collected from the qPCR-positive mice, and its seropositivity was verified for the particular strain used to infect that mouse using ELISA as described previously.^{79 (link)} Chondroitin lyase ABC from Proteus vulgaris was expressed in our laboratory. Recombinant Flavobacterial heparin lyases I, II, and III were expressed in our laboratory previously.^{80 (link)}