Beta-trace protein in ascites and pleural effusions: limits of CSF leakage detection.

Rhino- and/or otoliquorrhea can be diagnosed by detecting beta-trace protein (ß-TP) in nasal or ear secretions, as ß-TP is found in high concentrations in cerebrospinal fluid (CSF) but not in serum. CSF fistulae following trauma or surgery can also occur at other anatomical sites, resulting in CSF leakage into the thoracic and abdominal cavities. By analogy, determination of ß-TP has also been used to diagnose CSF admixture in pleural effusions and ascites. However, no systematic study has yet evaluated the concentrations of ß-TP in such fluids in the absence of CSF. To determine the validity of ß-TP determination as a marker for the presence of CSF, we investigated ß-TP concentrations in pleural effusions and ascites without CSF admixture. Patients from whom samples of ascites or pleural effusion and a paired plasma sample were available were investigated. One hundred sixty-four patients were prospectively recruited. ß-TP concentrations were determined by nephelometry. Mass spectrometric proteome analysis confirmed the presence of ß-TP in the samples. Median ß-TP concentrations detected in ascites and pleural effusions (range, 0.014-26.5 mg/L, median 2.29 mg/L) exceeded the corresponding plasma concentrations 2.6-fold. According to cutoffs published to diagnose rhino- and otoliquorrhea, between 6.1% and 95.7% of the specimens would have been erroneously rated CSF-positive. Protein analysis confirmed the presence of ß-TP in pleural effusion and ascites. Ascites and pleural effusion contain high concentrations of ß-TP that exceed the levels in corresponding plasma. Therefore, ß-TP is not a specific marker for the presence of CSF in these fluids.

Separation of beta2-transferrin by denaturing gel electrophoresis to detect cerebrospinal fluid in ear and nasal fluids.

BACKGROUND: Cerebrospinal fluid (CSF) leakage is a critical condition with a substantial risk of meningitis. We investigated the use of transferrin isoform analysis as a diagnostic marker for detection of CSF leakage in fluid samples. METHODS: We analyzed 241 samples from patients with CSF leakage, most commonly presenting as otorrhea or rhinorrhea, by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with subsequent Western blotting and immunostaining for transferrin. Tears, saliva, nasal fluid, and ear secretions (20 samples each) were analyzed in parallel, and normal human serum served as a control in each experiment. We compared the minimum volume of added CSF that could be detected in secretions by our assay with the minimum volume detected by the prostaglandin-D synthase (beta-trace) test. CSF was admixed with blood in different proportions to determine the influence of blood contamination on the transferrin pattern. RESULTS: In all CSF samples, beta1- and beta2-transferrin were present in nearly equal amounts. In tears and ear secretions, beta2-transferrin migrated in the gel in the same manner as in CSF, but its concentration was noticeably lower than that of beta1-transferrin, a difference that allowed a clear distinction from the transferrin pattern of CSF. In saliva, both transferrin isoforms were also present but could be distinguished from those of other fluids by electrophoretic migration pattern rather than relative concentrations. With the beta-trace test, a minimum of 5 microL of CSF was needed for detection, whereas our beta2-transferrin assay yielded a signal of comparable intensity with a minimum of 2 microL of CSF. CONCLUSION: Analysis of the transferrin microheterogeneity pattern by SDS-PAGE for the identification of CSF leakage is a highly sensitive and specific method that merits consideration as a routine technique.

Beta-trace protein as sensitive marker for CSF rhinorhea and CSF otorhea.

OBJECTIVE: Beta-trace protein concentrations in cerebrospinal fluid (CSF), serum and nasal secretions are investigated with a new quantitative, immunonephelometric assay. RESULTS: The mean beta-trace concentration of normal lumbar CSF (18.4 mg/l) and normal serum (0.59 mg/l), from n = 132 control patients, were 10% higher than reported earlier for smaller control groups. The reference range of beta-trace protein in nasal secretions is very low (median: 0.016 mg/l, range <0.003-0.12 mg/l, for n = 29 controls). Clinically confirmed cases of CSF rhinorhea (n = 20) showed beta-trace concentrations between 0.36 and 53.6 mg/l, with a median of 2.4 mg/l. We propose a cut-off value of 0.35 mg/l above which a CSF contamination in the secretion is plausible. A clinically confirmed CSF otorhea had a value of 1.75 mg/l. CONCLUSION: This new beta-trace protein assay offers a fast, sensitive and reliable routine method to detect a CSF rhinorhea or otorhea.

[Tuberculous otitis media. Report of 3 cases]. / Otitis media tuberculosa. Estudio de 3 casos.

Tuberculous otitis media (TOM) is a rare cause of chronic suppurative infection of the middle ear. Due to that the symptoms and signs are often indistinguishable from those of nontuberculosis chronic otitis media and the fact that the index of suspicion is low, there is frequently a considerable delay prior to diagnosis. This can lead to irreversible complications such as facial nerve paralysis and labyrinthitis. Medical therapy with antituberculous drugs is usually effective. We report three cases with TOM diagnosticated and followed up in our Service from january 1993 to july 2001. Their charts were retrospectively reviewed for relevant historical data, physical findings, complementary studies, treatment and clinical response. We performed a review of the literature, emphasizing that TOM should be considered in the differential diagnosis of chronic otitis media.

[Diagnosis and surgical treatment of liquorrhea]. / Diagnostikk og kirurgisk behandling av likvoré.

BACKGROUND: Liquorrhea caused by cerebrospinal fluid fistula may lead to meningitis and should be treated surgically. The diagnostic approach and the surgery may be difficult and improvements are needed. MATERIAL AND METHODS: We present two cases of liquorrhea. A review of the diagnostic and surgical methods is given. RESULTS: In both cases the diagnosis was confirmed by detection of beta-trace protein. In one case the surgery was complicated. This patient had a cerebrospinal fluid fistula from the sphenoid sinus. In a third operation, performed with sodium fluorescein dying, the fistula was closed. INTERPRETATION: Rhinorrhea and meningitis are unspecific signs of a possible cerebrospinal fluid fistula. Beta-trace protein is recommended as a diagnostic marker, because this protein has the highest positive predictive value for the presence of cerebrospinal fluid. High-resolution CT scans are the most helpful imaging technique for determining the leakage site, but this is not conclusive in all cases. Using sodium fluorescein may give better outcomes of surgical interventions.

[Analysis of beta-"trace" protein detects leakage of cerebrospinal fluid. Quickly, safely and cheaply]. / Analys av beta-"trace"-protein spårar läckage av cerebrospinalvätska. Snabbt, säkert och billigt.

beta-trace protein is an immunological marker for the detection of cerebrospinal fluid leakage to the nose or ear. Analysis of beta-trace protein via a nephelometric assay is a valuable test for the identification of cerebrospinal fluid in rhinorrhea or otorrhea.

Predictive values of beta-trace protein (prostaglandin D synthase) by use of laser-nephelometry assay for the identification of cerebrospinal fluid.

OBJECTIVE: beta-Trace protein (beta-TP) is an immunological marker for the detection of cerebrospinal fluid traces. The aim of the study was to evaluate the predictive values of a new research assay for beta-TP. METHODS: A total of 154 specimens from patients with otorrhea or rhinorrhea were investigated for cerebrospinal fluid (CSF) by use of a laser-nephelometric assay for beta-TP. Samples were obtained between January 1994 and November 2000. A sample was reported to be positive for CSF when the beta-TP concentration was more than 6 mg/L. Case evaluations were performed retrospectively and tabulated for indication, clinical course, additional investigations, surgical procedure, and follow-up. RESULTS: beta-TP was detected in 16 specimens; 138 samples were negative for beta-TP with a value less than 3 mg/L. One sample was suggestive of CSF traces at 4.6 mg/L. In correlation with the clinical course, the intraoperative findings, intraoperative visualization with sodium fluorescein, high-resolution computed tomography of the paranasal sinuses or the petrous bone, computed tomographic cisternography, magnetic resonance imaging, and radionuclide cisternography, there was no false-positive result. On four occasions, false-negative results occurred, with an overall accuracy of 0.974. The beta-TP test had a negative predictive value of 0.971 and a positive predictive value of 1. CONCLUSION: Analysis of betas-TP via the nephelometric assay is a valuable and reliable test in cranial base surgery for the identification of CSF.

A new nephelometric assay for beta-trace protein (prostaglandin D synthase) as an indicator of liquorrhoea.

OBJECTIVES: To determine the sensitivity and specificity of a nephelometric beta-trace protein assay for the diagnosis of liquorrhoea. METHODS: One hundred and forty clinical samples with suspected liquorrhoea were analysed by a newly developed nephelometric assay. An established electroimmunoassay served as a reference method. The sensitivity and specificity of the beta-trace nephelometric assay were calculated by a 2x2 contingency table for 10 different versions of a dichotomised nephelometric variable. In 52 patients (79 samples), the nephelometric findings were validated by referring to the clinical diagnosis based on the course of the disease, imaging techniques, and surgical inspection. RESULTS: Given a specificity of 100%, a beta-trace protein concentration of 6 mg/l or higher in a sample indicated liquorrhoea with a sensitivity of 92% compared with the reference method and of 93% compared with the clinical evaluation. The relation between the electroimmunoassay and the nephelometric assay was highly significant (p<0.001). CONCLUSIONS: The nephelometric beta-trace protein assay is a simple and rapid method for the detection of liquorrhoea with high sensitivity and specificity and may facilitate the diagnosis of fistulas leaking CSF.

[Beta-trace protein in diagnosis of cerebrospinal fluid fistula]. / Beta-trace-Protein in der Diagnostik der Liquorfistel.

Beta-trace protein is a lipocalin that was recently identified as prostaglandin D synthase and represents a major constituent of human cerebrospinal fluid. Beta-trace protein, similar to beta 2-transferrin, has been used as an immunological marker for the detection of cerebrospinal fluid. Between 1982 and 1999, 130 specimens from 101 patients with suspected cerebrospinal fluid leaks of the anterior or lateral skull base have been investigated for beta-trace protein. The specimens were analyzed by one-dimensional immunoelectrophoresis using Laurrell's electroimmunoassay. In all, beta-trace protein could be detected in 57 specimens and was absent in 73. All case notes were studied retrospectively. In correlation with the clinical course and the findings of CT of the paranasal sinuses, high resolution CT of the petrous bones, CT cisternography, NMR, 111In-DTPA radionuclide cisternography, intraoperative findings, visualization of sodium-fluorescein stained cerebrospinal fluid using endoscopic blue light or testing for glucose, there was no false-positive result. A false-negative result occurred in two cases. These findings show that the beta-trace test has a sensitivity of 97.3% and a specificity of nearly 100%. Our findings show that analysis of beta-trace protein can be a valuable test for detecting CSF leakage and has certain advantages in comparison with the beta 2-transferrin assay.

Quantification of beta-trace protein and detection of transferrin isoforms in mixtures of cerebrospinal fluid and blood serum as models of rhinorrhea and otorrhea diagnosis.

Different mixtures from a serum pool and a cerebrospinal fluid (CSF) pool were used as models to study CSF contamination in secretions by determining two CSF specific proteins: beta-trace protein (beta-TP) and the asialo-transferrin (a-Tf) band which was detected by isoelectric focusing (IEF) with Tf specific immunofixation. Beta-TP and total Tf were measured immunonephelometrically. Secretion/serum ratios of beta-TP content > 2.0 indicated CSF contaminations with > or = 5% (v/v) CSF; this was confirmed by detecting the a-Tf band by IEF. Reliable a-Tf bands were only revealed with secretion/serum rations of Tf contents < 0.1, indicating an interference of major sialo-Tf fractions with the a-TF band detection in the sample. For CSF detection in rhinorrhea and otorrhea, complementary use of beta-TP assay and a-Tf assay is recommended. Preanalytically, dilution or concentration of the sample as well as denaturation of Tf and beta-TP should be prevented by optimizing sample collection.

[The value of beta-2 transferrin analysis and MR cisternography for the diagnosis of cerebrospinal fluid fistulas]. / Intérêt de la recherche de beta2 transferrine et de la MR cisternographie pour le diagnostic des fistules de liquide céphalorachidien.

The diagnosis of cerebrospinal fluid (CSF) fistula may require invasive techniques. Detection of CSF and perilymph-specific beta 2 transferrin and MR cisternography which greatly enhances the CSF signal are sensitive and noninvasive techniques which allowed a precise diagnosis in seven patients with suspected CSF fistula. We review the different diagnostic techniques used for CSF fistula, beta 2 transferrin analysis and MR cisternography appear to provide accurate and noninvasive methods for investigating CSF fistulae. They should replace invasive techniques such as CT cisternography.

Detection of cerebrospinal fluid leakage by isoelectric focusing on polyacrylamide gels with silver staining using the PhastSystem.

Cerebrospinal fluid (CSF) leakage, which sometimes occurs after skull trauma, is a life-threatening condition. A prompt start with antibiotics and/or prompt surgical treatment of fistulas is essential to avoid severe complications. This requires a fast and reliable method for detecting CSF leakage. This paper describes a fast (< 2 h) method based on the identification of the tau protein (beta 2-transferrin) band(s). Tau protein is a brain-specific variant of transferrin that is characteristic of CSF. The method includes iso-electric focusing (IEF) on pre-cast polyacrylamide gels and silver staining using the PhastSystem, an automated instrument for electrophoresis and staining. In the present study, this technology was applied on 200 consecutive CSF samples, 32 of which were from healthy volunteers. Tau protein was detected in all CSF samples but 5 (2.5%), all of which were from patients with blood-brain barrier (BBB) damage. In these cases, the tau protein band was indistinct when direct silver staining was used. Therefore, immunofixation with an antitransferrin antibody was performed, and after that the tau protein band was easy to detect. The specificity of the method was high, since no brain-specific tau protein band was detected in serum, tears, saliva, or nasal secretion. As IEF of CSF using the PhastSystem is increasingly used as the routine method for detection of oligoclonal bands of IgG in neurological disorders, it could readily be used in the clinical (neuro) chemical laboratory also for the less frequent cases of suspected CSF leakage.

Detection of beta-2 transferrin in otorrhea and rhinorrhea in a routine clinical laboratory setting.

A simple, straightforward, and rapid method for the detection of beta-2 transferrin in otorrhea and rhinorrhea that can be used in a routine clinical laboratory is described. The beta-2 transferrin was detected by agarose gel electrophoresis of the fluid on Beckman Paragon equipment, followed by pressure transfer to a nitrocellulose membrane and then incubation with enzyme-labeled antitransferrin antibody and substrate. The procedure was fast (3.5 h) and sensitive (detected as little as 1 microgram/ml) and required only 3 microliters of fluid. Beta-2 transferrin was detected in cerebrospinal fluid diluted up to eightfold. No special training or expertise was needed, and all equipment and procedures used are commonly available in a routine clinical laboratory.

Beta-2 transferrin assay in clinical management of cerebral spinal fluid and perilymphatic fluid leaks.

Beta-2 transferrin is a protein found in cerebral spinal fluid (CSF) and inner ear perilymph, but not in blood, nasal or ear secretions. The purpose of this paper is to evaluate the clinical usefulness of the current assay for beta-2 transferrin for detecting CSF and perilymphatic leaks. We reviewed the hospital records of the first 88 patients having specimens submitted for beta-2 transferrin analysis at our institution. Both CSF and perilymph leaks were identified. However, confirmation of the absence or presence of beta-2 transferrin was directly used in the clinical management of only 55% of the patients. This was largely secondary to the time delay in test processing and initial lack of physician confidence with the test. However, our review of the clinical outcomes relating to the use of the beta-2 transferrin analysis suggests high sensitivity and specificity for the test. Analysis of beta-2 transferrin appears to be a valuable test for detecting CSF leakage and a promising test for confirming perilymphatic leaks. However, to achieve greater clinical usefulness a rapid clinical assay needs to be developed and further information gained regarding the sensitivity and specificity of the beta-2 transferrin assay for detecting CSF and perilymphatic fluid leakage.

The concept of transtympanic injection of fibrin caulk.

Cerebrospinal fluid otorhinorrhea after basilar skull trauma poses a difficult management problem. When conservative techniques fail, more aggressive neurosurgical and otologic procedures are required to control cerebrospinal fluid leakage. We assessed a less invasive method for the repair of traumatic cerebrospinal fluid fistulas. Thirty-one adult Sprague-Dawley rats were used to develop an animal model for the treatment of cerebrospinal fluid leakage. A fistula was created by removing a thin plate of bone from the superior aspect of the rat bulla. The bulla was then plugged with a transtympanic injection of fibrin caulk. Otoscopic and histologic data were collected at selected intervals. Transtympanic injection of fibrin caulk failed to alter significantly the rate of healing of cerebrospinal fluid fistulas. Coagulum retraction, rapid fibrinolysis, and other reasons for failure are explored.

Highly sensitive detection of beta-2 transferrin in rhinorrhea and otorrhea as a marker for cerebrospinal fluid (C.S.F.) leakage.

As beta-2 transferrin is almost specific to the cerebrospinal fluid, its detection in rhinorrhea or otorrhea fluids demonstrates the occurrence of CSF fistula. We describe a highly sensitive method based on immunoaffinity-mediated capillary blotting for the detection of this cathodic isoform of transferrin in minute amounts (3 microliters) of rhinorrhea or otorrhea fluids. Application of this method in a series of 10 patients with CSF fistula is reported.

[Cerebrospinal otorrhea--cerebrospinal rhinorrhea. The Salzburg concept of cerebrospinal fluid diagnosis]. / Otoliquorrhoe--Rhinoliquorrhoe. Salzburger Konzept zur Liquordiagnostik.

The three following cerebrospinal fluid (CSF) examinations make it possible to identify even the smallest amounts of CSF in case of CSF otorrhoea and rhinorrhoea. 1. Immunological identification of beta 2-transferrin (Oberascher/Arrer) 2. Laboratory fluorescein identification (Oberascher/Arrer) 3. Endoscopic fluorescein detection according to Messerklinger. For screening, and as the method of choice, beta 2-transferrin identification is always used as a first step if there is a suspicion of liquorrhoea. Depending on the result and on further measures, both fluorescein tests are used additionally in diagnosis. Basing on practical experience gained recently, special attention is given to test analysis, the various possibilities of taking samples, and their means of transport or mailing. A newly developed diagnostic step-by-step plan is intended to emphasise the clinical significance by means of practical examples. This concept represents the present state of the art in CSF diagnosis and demonstrates that a much mor precise range of indication is possible in surgery of fractures of the base of the skull and CSF leaks if it is combined with an appropriate x-ray examination.

Cerebrospinal fluid otorrhea--new trends in diagnosis.

In this paper we describe the different causes of cerebrospinal fluid (CSF) otorrhea and their pathomechanisms, followed by a short description and illustration of the most common methods used until now for identifying CSF. We then present a new and modern method, immunologic identification of CSF using beta 2-transferrin (tau band). This protein variant is found only in CSF, not in any other body fluids, such as tears, nasal secretions, saliva, or blood serum. Analysis of CSF using this method always indicates two bands, the beta 1-transferrin and the beta 2-transferrin band. The latter is typical for CSF. The analysis of all other body fluids shows just one band, the beta 1-transferrin band. It is therefore possible to identify CSF accurately. The required reagents and equipment are listed and methods of taking samples are explained, followed by a detailed description of sample preparation, electrophoresis, immunofixation, and silver staining. Staining with alkaline silver nitrate has a 40-fold higher sensitivity than staining with the commonly used coomassie brilliant blue. With this method, 1 microliter pure CSF (corresponding to approximately 1/50 of a drop) and 100 microliter CSF (two drops) per 1 ml wound secretion can be identified.

Beta trace-protein as marker for cerebrospinal fluid fistula.

The detection of beta trace-protein with the help of an easy-to-perform Ouchterlony test is strongly indicative of liquorrhea. The protein has been purified from pooled cerebrospinal fluid and an antiserum has been produced in rabbits. An open CSF fistula was diagnosed in 24 cases and there was no false positive result.