The Laryngoscope V C 2014 The American Laryngological, Rhinological and Otological Society, Inc. False Negative b-2 Transferrin in the Diagnosis of Cerebrospinal Fluid Leak in the Presence of Streptococcus pneumoniae Maya Korem, MD; Haim Ovadia, PhD; Iddo Paldor, MD; Allon E. Moses, MD; Colin Block, MD; Ron Eliashar, MD; Nir Hirshoren, MD Objectives/Hypothesis: The objectives of this study were to examine the presence of b-2 transferrin (b2TRNSF) in cer- ebrospinal fluid (CSF) contaminated in vitro by various bacteria and explore the mechanism (passive or active) responsible for b2TRNSF elimination. Early diagnosis of CSF leakage may change treatment decisions and minimize the risk of meningitis and encephalitis. b2TRNSF is a protein present exclusively in CSF. Its detection is highly useful in cases of CSF leakage, although it has never been examined in the presence of central nervous system infection. Study Design: Prospective patient analysis. Methods: Sterile CSF drawn from patients was contaminated in vitro with several microorganisms chosen for their abil- ity to cause neurosurgical-related infections: Streptococcus pneumoniae, methicillin-sensitive Staphylococcus aureus, Staphylo- coccus epidermidis, and Pseudomonas aeruginosa. b2TRNSF was examined at two time points: following immediate inoculation (t 0 ) and following an overnight incubation (t 18 ) over various bacterial concentrations. Samples of CSF inoculated with S pneu- moniae were also examined in the presence of ciprofloxacin. For b2TRNSF analysis we used immunoblotting electrophoresis and enzyme-linked immunosorbent assay (ELISA). Results: CSF samples collected from nine patients were analyzed. b2TRNSF was not detected following S pneumoniae inoculation at both time points when immunoblotting electrophoresis was used. Quantitative analysis using ELISA demon- strated significant b2TRNSF concentration decrease. The addition of ciprofloxacin led to the same results. Conclusions: CSF leak detection using b2TRNSF may be deceiving in the presence of a S pneumoniae cerebral nervous system infection. A passive process is suggested, as b2TRNSF disappeared either immediately or following incubation with inactive bacteria. Key Words: b-2 transferrin, cerebrospinal fluid leakage, electrophoresis, enzyme-linked immunosorbent assay, Strepto- coccus pneumoniae. Level of Evidence: NA Laryngoscope, 125:556–560, 2015 INTRODUCTION Cerebrospinal fluid (CSF) leakage secondary to fis- tula formation between the dura matter and skull base may result from traumatic head injury, iatrogenic injury, or spontaneously (associated with low or high intracra- nial pressure, as in congenital anomalies, brain tumors, cysts, and hydrocephalus). 1–4 Free passage of bacterial flora from the nasal cavity and paranasal sinuses through a CSF fistula into the cranium may pose a risk for meningitis and encephalitis. Meningitis may super- vene in as many as 19% of such cases. 5 Thus, early diag- nosis and treatment of CSF leakage may minimize the risk of a lethal infection. The b-2 transferrin (b2TRNSF) protein was intro- duced as a marker for CSF by Meurman et al. in 1979. 6 b2TRNSF detection is a highly reliable sensitive diagnos- tic method for detecting CSF leakage. 7,8 This glycoprotein is a desialylated isoform of transferrin without neura- minic acid and is found almost exclusively in the CSF. It is believed that b2TRNSF is converted from the b-1 iso- form via cerebral neuraminidase rather than produced de novo. 9 Therefore, the presence of b2TRNSF may be used as a marker for CSF leakage even when the CSF is mixed with blood or with other secretions. 1,10–12 A common laboratory technique for b2TRNSF detec- tion is immunofixation electrophoresis. The presence of two transferrin bands in b-globulin fraction indicates a positive result for CSF. To achieve good sensitivity and compensate for a low CSF b2TRNSF concentration, 2 to 5 mL are required, 13 whereas only 2 lL 10 are required if immunoblotting electrophoresis is used. The ability to detect b2TRNSF in CSF samples of patients suffering from bacterial central nervous system (CNS) infections has never been explored. We had two patients with pneumococcal meningitis and clinical CSF From the Department of Clinical Microbiology and Infectious Dis- eases (M.K., A.E.M., C.B.), Department of Neurology (H.O.), the Agnes Ginges Center for Human Neurogenetics, Department of Neurosurgery (I.P .), and the Department of Otolaryngology/Head & Neck Surgery (R.E., N.H.), Hebrew University School of Medicine–Hadassah Medical Center, Jerusalem, Israel. Editor’s Note: This Manuscript was accepted for publication August 29, 2014. This study was supported by the Hebrew University School of Medicine–Hadassah Medical Center Grant for Young Clinician (N.H.). The authors have no other funding, financial relationships, or con- flicts of interest to disclose. Send correspondence to Nir Hirshoren, MD, Department of Otolar- yngology/Head & Neck Surgery, Hadassah Ein-Kerem, Jerusalem, 91120, Israel. E-mail: drnir@hadassah.org.il DOI: 10.1002/lary.24940 Laryngoscope 125: March 2015 Korem et al.: Misdiagnosis of Cerebrospinal Fluid Leak 556