The potential mechanisms for blood-brain barrier damage and the diagnosis of neurosyphilis in HIV patients co-infected with syphilis (HIV-S) are unclear. The aim of the study was to determine the expression of CXCL2 in the serum and cerebrospinal fluid (CSF) of HIV-S patients. A total of 34 HIV patients and 7 controls were enrolled in a HIV clinical cohort for diagnosis of neurosyphilis in Taiwan. Serum and CSF concentrations of CXCL2 were determined by ELISA. Neurosyphilis was defined as a CSF white blood cell count of ≧20 cells/μl or a reactive CSF Venereal Disease Research Laboratory (VDRL). Demographics and medical histories were collected. All the patients with HIV-S were males. Most (80%) had sex with men (MSM) and serum rapid plasma reagin (RPR) titers of ≧1:32. The medium age was 37 (range 21-68) years. The medium CD4 T cell counts at the time of the diagnosis of syphilis were 299 (range 92-434) cells/μl. Eight patients (24%) had neurosyphilis based on a reactive CSF VDRL test (n = 5) or increased CSF white blood cell counts of ≧20 cells/μl (n = 3). The concentrations of CSF CXCL2 were significantly higher in patients with HIV and neurosyphilis as compared to HIV with syphilis, HIV, and controls (p = 0.012). There were no significant differences in serum concentrations between the four groups. There was a correlation between CSF CXCL2 concentrations with neurosyphilis (p = 0.017), CSF white blood cell count (p = 0.001), and CSF protein levels (p = 0.005). The CSF level of CXCL2 can be used to distinguish those with or without neurosyphilis in HIV infected patients.