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Background: Cerebrospinal fluid (CSF) leaks can lead to serious complications if left untreated, making rapid and accurate diagnosis essential. Biomarkers such as β2-transferrin (B2TRF) and β-trace protein are used to detect CSF leaks, but their limitations warrant the exploration of alternative markers. This study investigates the potential of phosphorylated tau at threonine 181 (p-tau 181 ) as a biomarker for CSF leaks., Methods: Samples from 56 subjects were analyzed for B2TRF and p-tau 181 using immunoaffinity blotting and chemiluminescent enzyme immunoassay, respectively. Data analysis included Mann-Whitney test to assess the overall difference in median p-tau 181 concentrations between B2TRF positive and negative patients and a receiver operating characteristic (ROC) curve analysis to determine optimal p-tau 181 cutoff values for predicting B2TRF positivity., Results: p-tau 181 levels were significantly higher in B2TRF positive samples compared to negative samples (p < 0.001). ROC analysis showed high diagnostic performance for p-tau 181 , with an optimal cutoff of 13.22 pg/mL providing 92.0% sensitivity and 93.1% specificity. Excluding hemolyzed samples improved further the diagnostic performances, maintaining high sensitivity (90.9%) and achieving perfect specificity (100.0%)., Conclusions: This study highlights the potential of p-tau 181 as a valuable biomarker for the detection of CSF leaks due to its high diagnostic accuracy and practical advantages over the current biomarkers. The characteristics of p-tau 181 assay being both quantitative and rapid, with high diagnostic accuracy, suggest that it could be a valuable tool for the detection of CSF leaks. Further research are now needed to validate these findings., (© 2024. Springer-Verlag GmbH Germany, part of Springer Nature.)

Background and Objectives: Rapid detection of cerebrospinal fluid (CSF) leaks is vital for patient recovery after spinal surgery. However, distinguishing CSF-specific transferrin (TF) from serum TF using lateral flow immunoassays (LFI) is challenging due to their structural similarities. This study aims to develop a novel point-of-care diagnostic assay for precise CSF leak detection by quantifying total TF in both CSF and serum., Methods: Capitalizing on the substantial 100-fold difference in TF concentrations between CSF and serum, we designed a diagnostic platform based on the well-known "hook effect" resulting from excessive analyte presence. Clinical samples from 37 patients were meticulously tested using the novel LFI sensor, alongside immunofixation as a reference standard., Results: The hook effect-based LFI sensor exhibited outstanding performance, successfully discriminating positive clinical CSF samples from negative ones with remarkable statistical significance (positive vs negative t -test; P = 1.36E-05). This novel sensor achieved an impressive 100% sensitivity and 100% specificity in CSF leak detection, demonstrating its robust diagnostic capabilities., Conclusion: In conclusion, our study introduces a rapid, highly specific, and sensitive point-of-care test for CSF leak detection, harnessing the distinctive TF concentration profile in CSF compared with serum. This novel hook effect-based LFI sensor holds great promise for improving patient outcomes in the context of spinal surgery and postsurgical recovery. Its ease of use and reliability make it a valuable tool in clinical practice, ensuring timely and accurate CSF leak detection to enhance patient care., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the Congress of Neurological Surgeons.)

Background: Beta-2 transferrin (B2-Tf) gel electrophoresis (GE) is the preferred non-invasive diagnostic modality for confirming cerebrospinal fluid (CSF) in body fluids. While B2-Tf GE testing is highly sensitive and specific for CSF, false-positive (FP) and false-negative (FN) results can lead to diagnostic and therapeutic dilemmas. Several series have demonstrated potential causes of false B2-Tf GE results, but few studies have reported reasons for these errors. The purpose of this systematic review was to describe sources of B2-Tf GE errors., Methods: A systematic review was performed by searching OVID, EMBASE, and Web of Science databases for B2-Tf GE studies. After applying exclusion criteria, original research studies directly addressing erroneous B2-Tf GE results underwent qualitative analysis., Results: Of the 243 abstracts screened, 71 underwent full-text review and 18 studies reporting B2-Tf GE errors were included for analysis. There were 15 potential FPs, 12 actual FPs, 12 potential FNs, 19 actual FNs, and 14 indeterminate results. There were also 246 potentially indeterminate results from in vitro studies. Reasons for B2-Tf GE errors included serum transferrin alterations (n = 17; all potential), infection related (n = 13; 9 potential), orbital or salivary contamination (n = 2; 1 potential), and collection related (n = 255; 246 potential). There were 31 false or indeterminate results with unspecified reasons. There were no reported errors due to laboratory processing., Conclusions: Multiple potential or actual reasons for false or indeterminate results have been reported for B2-Tf GE testing of rhinorrhea and otorrhea. Future studies should explore reasons for B2-Tf testing errors and how these may affect clinical decision making., (© 2023 ARS‐AAOA, LLC.)

Cerebrospinal fluid (CSF) leakage may lead to life-threatening complications if not detected promptly. However, gel electrophoresis, the gold-standard test for confirming CSF leakage by detecting beta2-transferrin (β2-Tf), requires 3-6 h and is labor-intensive. We developed a new β2-Tf detection platform for rapid identification of CSF leakage. The three-step design, which includes two steps of affinity chromatography and a rapid sensing step using a semiconductor-enriched single-walled carbon nanotube field-effect transistor (FET) sensor, circumvented the lack of selectivity that antitransferrin antibody exhibits for transferrin isoforms and markedly shortened the detection time. Furthermore, three different sensing configurations for the FET sensor were investigated for obtaining the optimal β2-Tf sensing results. Finally, body fluid (CSF and serum) tests employing our three-step strategy demonstrated high sensitivity, suggesting its potential to be used as a rapid diagnostic tool for CSF leakage.

HIV-associated neurocognitive disorder (HAND) remains prevalent despite antiretroviral therapy and involves white matter damage in the brain. Although iron is essential for myelination and myelin maintenance/repair, its role in HAND is largely unexplored. We tested the hypotheses that cerebrospinal fluid (CSF) heavy-chain ferritin (Fth1) and transferrin, proteins integral to iron delivery and myelination, are associated with neurocognitive performance in people with HIV (PWH). Fth1, transferrin, and the pro-inflammatory cytokines TNF-α and IL-6 were quantified in CSF at baseline (entry) in 403 PWH from a prospective observational study who underwent serial, comprehensive neurocognitive assessments. Associations of Fth1 and transferrin with Global Deficit Score (GDS)-defined neurocognitive performance at baseline and 30-42 months of follow-up were evaluated by multivariable regression. While not associated with neurocognitive performance at baseline, higher baseline CSF Fth1 predicted significantly better neurocognitive performance over 30 months in all PWH (p < 0.05), in PWH aged < 50 at 30, 36, and 42 months (all p < 0.05), and in virally suppressed PWH at all three visit time-points (all p < 0.01). Higher CSF transferrin was associated with superior neurocognitive performance at all visits, primarily in viremic individuals (all p < 0.05). All associations persisted after adjustment for neuro-inflammation. In summary, higher CSF Fth1 is neuroprotective over prolonged follow-up in all and virally suppressed PWH, while higher CSF transferrin may be most neuroprotective during viremia. We speculate that higher CSF levels of these critical iron-delivery proteins support improved myelination and consequently, neurocognitive performance in PWH, providing a rationale for investigating their role in interventions to prevent and/or treat HAND., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Objective: Iron has been implicated in the pathogenesis of brain injury and hydrocephalus after preterm germinal matrix hemorrhage-intraventricular hemorrhage, however, it is unknown how external or endogenous intraventricular clearance of iron pathway proteins affect the outcome in this group., Methods: This prospective multicenter cohort included patients with posthemorrhagic hydrocephalus (PHH) who underwent (1) temporary and permanent cerebrospinal fluid (CSF) diversion and (2) Bayley Scales of Infant Development-III testing around 2 years of age. CSF proteins in the iron handling pathway were analyzed longitudinally and compared to ventricle size and neurodevelopmental outcomes., Results: Thirty-seven patients met inclusion criteria with a median estimated gestational age at birth of 25 weeks; 65% were boys. Ventricular CSF levels of hemoglobin, iron, total bilirubin, and ferritin decreased between temporary and permanent CSF diversion with no change in CSF levels of ceruloplasmin, transferrin, haptoglobin, and hepcidin. There was an increase in CSF hemopexin during this interval. Larger ventricle size at permanent CSF diversion was associated with elevated CSF ferritin (p = 0.015) and decreased CSF hemopexin (p = 0.007). CSF levels of proteins at temporary CSF diversion were not associated with outcome, however, higher CSF transferrin at permanent CSF diversion was associated with improved cognitive outcome (p = 0.015). Importantly, longitudinal change in CSF iron pathway proteins, ferritin (decrease), and transferrin (increase) were associated with improved cognitive (p = 0.04) and motor (p = 0.03) scores and improved cognitive (p = 0.04), language (p = 0.035), and motor (p = 0.008) scores, respectively., Interpretation: Longitudinal changes in CSF transferrin (increase) and ferritin (decrease) are associated with improved neurodevelopmental outcomes in neonatal PHH, with implications for understanding the pathogenesis of poor outcomes in PHH. ANN NEUROL 2021;90:217-226., (© 2021 American Neurological Association.)

Proper functioning of all organs, including the brain, requires iron. It is present in different forms in biological fluids, and alterations in its distribution can induce oxidative stress and neurodegeneration. However, the clinical parameters normally used for monitoring iron concentration in biological fluids (i.e., serum and cerebrospinal fluid) can hardly detect the quantity of circulating iron, while indirect measurements, e.g., magnetic resonance imaging, require further validation. This review summarizes the mechanisms involved in brain iron metabolism, homeostasis, and iron imbalance caused by alterations detectable by standard and non-standard indicators of iron status. These indicators for iron transport, storage, and metabolism can help to understand which biomarkers can better detect iron imbalances responsible for neurodegenerative diseases.

Background: Iron plays an important role in maintaining cell survival, with normal iron trafficking known to be regulated by the ceruloplasmin-transferrin (Cp-Tf) antioxidant system. Disruption to this system is thought to be detrimental to normal brain function., Objective: To determine whether an imbalance of iron and the proteins involved in its metabolism (ceruloplasmin and transferrin) are linked to Alzheimer's disease (AD) and to the expression of amyloid-beta (Aβ) peptide 1-42 (Aβ1-42), which is a major species of Aβ, and the most toxic., Methods: We evaluated the concentrations of iron, calcium, magnesium, and Aβ1-42 in the cerebrospinal fluid (CSF) of patients with AD and cognitively normal controls. Correlations between the components of the Cp-Tf antioxidant system in plasma were studied to determine the role of peripheral blood in the onset and/or development of AD. We used commercial ELISA immunoassays to measure Aβ1-42, immunoturbidimetry to quantify ceruloplasmin and transferrin, and colorimetry to quantify iron, calcium, and magnesium., Results: We found that the AD group had lower CSF concentrations of Aβ1-42 (p < 0.001) and calcium (p < 0.001), but a higher CSF concentration of iron (p < 0.001). Significantly lower plasma concentrations of ceruloplasmin (p = 0.003), transferrin (mean, p < 0.001), and iron (p < 0.001) were observed in the AD group than in cognitively normal adults. Moreover, we found a strong interdependence between most of these components., Conclusion: Iron dyshomeostasis has a crucial role in the onset of AD and/or its development. Correcting metal misdistribution is an appealing therapeutic strategy for AD.

Background: Iron is crucial for proper functioning of all organs including the brain. Deficiencies and excess of iron are common and contribute to substantial morbidity and mortality. Whereas iron's involvement in erythropoiesis drives clinical practice, the guidelines informing interventional strategies for iron repletion in neurological disorders are poorly defined. The objective of this study was to determine if peripheral iron status is communicated to the brain., Methods: We used a bi-chamber cell culture model of the blood-brain-barrier to determine transcytosis of iron delivered by transferrin as a metric of iron transport. In the apical chamber (representative of the blood) we placed transferrin complexed with iron 59 and in the basal chamber (representative of the brain) we placed human cerebrospinal fluid. Cerebrospinal fluid (CSF) samples (N = 24) were collected via lumbar puncture. The integrity of the tight junctions were monitored throughout the experiments using RITC-Dextran., Results: We demonstrate that iron transport correlates positively with plasma hemoglobin concentrations but not serum ferritin levels., Conclusions: The clinical ramifications of these findings are several- fold. They suggest that erythropoietic demands for iron take precedence over brain requirements, and that the metric traditionally considered to be the most specific test reflecting total body iron stores and relied upon to inform treatment decisions-i.e., serum ferritin-may not be the preferred peripheral indicator when attempting to promote brain iron uptake. The future direction of this line of investigation is to identify the factor(s) in the CSF that influence iron transport at the level of the BBB.

Objective: The objective of this study was to compare iron metabolic variables in the serum and cerebrospinal fluid (CSF) of patients with sporadic amyotrophic lateral sclerosis (sALS) with those of patients with multiple system atrophy (MSA) and control subjects. We also assessed the correlations of these variables with sALS progression and severity and estimated their roles in predicting prognosis. Methods: We retrospectively collected iron metabolic parameters, including serum levels of iron, ferritin, transferrin levels and total iron binding capacity and the CSF level of ferritin, from 435 sALS patients, 176 MSA patients and 431 control subjects. Results: Serum ferritin levels were significantly higher in the sALS group compared with the MSA and control groups in both males ( p  = 0.001 and p  < 0.0001, respectively) and females ( p  = 0.034 and p  < 0.0001, respectively). However, serum transferrin levels were significantly lower in females of the sALS group compared with the MSA ( p  = 0.016) and control ( p  = 0.015) groups. The CSF ferritin level and the serum levels of total iron binding capacity and iron were similar among the sALS, MSA and control groups. Survival analysis demonstrated that higher serum ferritin levels were predictors of reduced survival of sALS patients. No correlations between iron metabolic variables and clinical parameters were found. Conclusion: An elevated serum ferritin level is associated with reduced survival of sALS patients. However, the levels of iron metabolic parameters were not associated with clinical deterioration or disease severity at diagnosis.

Iron plays many important roles in the brain, including involvement in myelination, neurotransmission and electron transfer in the respiratory chain. Transferrin (Tf), an iron transporter, is mainly biosynthesized in the liver, but can also be biosynthesized in the brain; i.e., by oligodendrocytes and the choroid plexus, a cerebrospinal fluid (CSF) producing tissue. The CSF contains two Tf isoforms, brain-type Tf and serum-type Tf, which differ in their glycan structures. Brain-type Tf is uniquely glycolsylated with biantennary asialo- and agalacto-complex type N-glycans that carry bisecting β1,4-GlcNAc and core α1,6-Fuc. The glycans of serum-type Tf in the CSF are similar to those of Tf in serum. Biochemical analyses reveal that the apparent molecular size of brain-type Tf is smaller than that of serum-type Tf, and that hydrophobic patches are exposed on brain-type Tf as demonstrated by hydrophobic probe binding studies. We found that brain-type Tf levels were decreased in idiopathic normal pressure hydrocephalus, in which CSF production is suspected to decrease, while brain-type Tf increased in spontaneous intracranial hypotension, in which CSF production is suspected to increase. These results suggest that brain-type Tf could be a biomarker of altered CSF production.

Idiopathic normal pressure hydrocephalus (iNPH) is a dementia-inducing disorder. Primary cause of iNPH is speculated to be a reduction of cerebrospinal fluid (CSF) absorption, which secondarily induces hydrocephalus, compression of brain, and reduction of CSF production. Patients are treated by surgically inserting a shunt to deliver excess CSF to the abdominal cavity. The prognosis for cognitive improvement after shunt surgery has been difficult to predict. We therefore investigated various CSF proteins, hoping to find a biomarker predictive of cognitive performance one to two years after shunt surgery. CSF proteins of 34 iNPH and 15 non-iNPH patients were analysed by Western blotting, revealing two glycan isoforms of transferrin (Tf); 'brain-type' Tf with N-acetylglucosaminylated glycans and 'serum-type' Tf with α2, 6-sialylated glycans. Brain-type Tf levels decreased in iNPH but rapidly returned to normal levels within 1-3 months after shunt surgery. This change was positively correlated with recovery from dementia, per Mini-Mental State Examination and Frontal Assessment Battery scores at 11.8 ± 7.7 months post-operation, suggesting that brain-type Tf is a prognostic marker for recovery from dementia after shunt surgery for iNPH. Histochemical staining with anti-Tf antibody and an N-acetylglucosamine-binding lectin suggests that brain-type Tf is secreted from choroid plexus, CSF-producing tissue.

Background: The effect of time and temperature on beta-2 transferrin stability in cerebrospinal fluid (CSF) is not well established. After collecting nasal CSF for testing, beta-2 transferrin has been found to be stable and detectable for 1 week, whether being refrigerated or stored at room temperature. The purpose of this study was to determine if beta-2 transferrin remained detectable longer than 1 week and whether refrigeration improved its detectability., Methods: In patients undergoing therapeutic CSF diversion, 2-mL CSF samples were collected from 18 patients. The samples were divided and stored either at room temperature, or at 4°C, and tested for beta-2 transferrin at 7 and 14 days. CSF was collected from external ventricular drains (EVDs) (n = 15), lumbar drains (n = 2), and subdural drains (n = 1)., Results: Of the 18 CSF samples originally testing positive for beta-2 transferrin, none turned negative at 7 or 14 days, in both the refrigerated and room temperature groups (95% confidence interval [CI], 0% to 18.5%)., Conclusion: Beta-2 transferrin remained detectable for 14 days in all CSF samples, regardless of being stored at 4°C or room temperature., (© 2018 ARS-AAOA, LLC.)

Background: Spontaneous intracranial hypotension (SIH) is caused by cerebrospinal fluid (CSF) leakage. Definitive diagnosis can be difficult by clinical examinations and imaging studies., Methods: SIH was diagnosed with the following criteria: (i) evidence of CSF leakage by cranial magnetic resonance imaging (MRI) findings of intracranial hypotension and/or low CSF opening pressure; (ii) no recent history of dural puncture. We quantified CSF proteins by ELISA or Western blotting., Results: Comparing with non-SIH patients, SIH patients showed significant increase of brain-derived CSF glycoproteins such as lipocalin-type prostaglandin D synthase (L-PGDS), soluble protein fragments generated from amyloid precursor protein (sAPP) and "brain-type" transferrin (Tf). Serum-derived proteins such as albumin, immunoglobulin G, and serum Tf were also increased. A combination of L-PGDS and brain-type Tf differentiated SIH from non-SIH with sensitivity 94.7% and specificity 72.6%., Conclusion: L-PGDS and brain-type Tf can be biomarkers for diagnosing SIH., General Significance: L-PGDS and brain-type Tf biosynthesized in the brain appears to be markers for abnormal metabolism of CSF., (Copyright © 2018. Published by Elsevier B.V.)

Objectives: Iron deficiency (ID) anemia leads to long-term neurodevelopmental deficits by altering iron-dependent brain metabolism. The objective of the study was to determine if ID induces metabolomic abnormalities in the cerebrospinal fluid (CSF) in the pre-anemic stage and to ascertain the aspects of abnormal brain metabolism affected., Methods: Standard hematological parameters [hemoglobin (Hgb), mean corpuscular volume (MCV), transferrin (Tf) saturation, and zinc protoporphyrin/heme (ZnPP/H)] were compared at 2, 4, 6, 8, and 12 months in iron-sufficient (IS; n = 7) and iron-deficient (ID; n = 7) infant rhesus monkeys. Five CSF metabolite ratios were determined at 4, 8, and 12 months using 1 H NMR spectroscopy at 16.4 T and compared between groups and in relation to hematologic parameters., Results: ID infants developed ID (Tf saturation < 25%) by 4 months of age and all became anemic (Hgb < 110 g/L and MCV < 60 fL) at 6 months. Their heme indices normalized by 12 months. Pyruvate/glutamine and phosphocreatine/creatine (PCr/Cr) ratios in CSF were lower in the ID infants by 4 months (P < 0.05). The PCr/Cr ratio remained lower at 8 months (P = 0.02). ZnPP/H, an established blood marker of pre-anemic ID, was positively correlated with the CSF citrate/glutamine ratio (marginal correlation, 0.34; P < 0.001; family wise error rate = 0.001)., Discussion: Metabolomic analysis of the CSF is sensitive for detecting the effects of pre-anemic ID on brain energy metabolism. Persistence of a lower PCr/Cr ratio at 8 months, even as hematological measures demonstrated recovery from anemia, indicate that the restoration of brain energy metabolism is delayed. Metabolomic platforms offer a useful tool for early detection of the impact of ID on brain metabolism in infants.

Background: Cerebrospinal fluid (CSF) is sequestered from blood by the blood-brain barrier and directly communicates with brain parenchymal interstitial fluid, leading to contain specific biomarkers of neurological diseases., Scope of Review: CSF contains glycan isoforms of transferrin (Tf): one appears to be derived from the brain and the other from blood., Major Conclusions: CSF contains two glycan-isoforms; brain-type Tf and serum-type Tf. Glycan analysis and immunohistochemistry suggest that serum-type Tf having α2, 6sialylated glycans is derived from blood whereas brain-type Tf having GlcNAc-terminated glycans is derived from the choroid plexus, CSF producing tissue. The ratio of serum-type/brain-type Tf differentiates Alzheimer's disease from idiopathic normal pressure hydrocephalus, which is an elderly dementia caused by abnormal metabolism of CSF. The ratios in Parkinson's disease (PD) patients were higher than those of controls and did not appear to be normally distributed. Indeed, detrended normal Quantile-Quantile plot analysis reveals the presence of an independent subgroup showing higher ratios in PD patients. The subgroup of PD shows higher levels of CSF α-synuclein than the rest, indicating that PD includes two subgroups, which differ in levels of brain-type Tf and α-synuclein., General Significance: Glycosylation in central nervous system appears to be unique. The unique glycan may be a tag for glycoprotein, which is biosynthesized in the central nervous system. This article is part of a Special Issue entitled Neuro-glycoscience, edited by Kenji Kadomatsu and Hiroshi Kitagawa., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Background: HIV-associated neurocognitive disorder (HAND) remains common, despite antiretroviral therapy (ART). HIV dysregulates iron metabolism, but cerebrospinal fluid (CSF) levels of iron and iron-transport proteins in HIV-infected (HIV+) persons are largely unknown. The objectives of this study were to characterize CSF iron-related biomarkers in HIV+ adults and explore their relationships to known predictors of HAND., Methods: We quantified total iron, transferrin and heavy-chain (H)-ferritin by immunoassay in CSF sampled by lumbar puncture in 403 HIV+ participants in a multi-center, observational study and evaluated biomarker associations with demographic and HIV-related correlates of HAND [e.g., age, sex, self-reported race/ethnicity, ART, and detectable plasma virus and CSF viral load (VL)] by multivariable regression. In a subset (N = 110) with existing CSF: serum albumin (Q Alb ) measurements, Q Alb and comorbidity severity were also included as covariates to account for variability in the blood-CSF-barrier., Results: Among 403 individuals (median age 43 years, 19% women, 56% non-Whites, median nadir CD4+ T cell count 180 cells/µL, 46% with undetectable plasma virus), men had 25% higher CSF transferrin (median 18.1 vs. 14.5 µg/mL), and 71% higher H-ferritin (median 2.9 vs. 1.7 ng/mL) than women (both p-values ≤0.01). CSF iron was 41% higher in self-reported Hispanics and 27% higher in (non-Hispanic) Whites than in (non-Hispanic) Blacks (median 5.2 and 4.7 µg/dL in Hispanics and Whites, respectively, vs. 3.7 µg/dL in Blacks, both p ≤ 0.01); these findings persisted after adjustment for age, sex, and HIV-specific factors. Median H-ferritin was 25% higher (p < 0.05), and transferrin 14% higher (p = 0.06), in Whites than Blacks. Transferrin and H-ferritin were 33 and 50% higher, respectively, in older (age > 50 years) than in younger persons (age ≤ 35 years; both p < 0.01), but these findings lost statistical significance in subset analyses that adjusted for Q Alb and comorbidity. After these additional adjustments, associations were observed for CSF iron and transferrin with race/ethnicity as well as CSF VL, for transferrin with sex and ART, and for H-ferritin with plasma virus detectability and significant comorbidity (all p < 0.05)., Conclusions: CSF iron biomarkers are associated with demographic factors, ART, and CSF VL in HIV+ adults. Future studies should investigate a role for CNS iron dysregulation, to which an altered blood-CSF barrier may contribute, in HAND.

Glycosylation is one of the most common and important post-translational modifications, influencing both the chemical and the biological properties of proteins. Studying the glycosylation of the entire protein population of a sample can be challenging because variations in the concentrations of certain proteins can enhance or obscure changes in glycosylation. Furthermore, alterations in the glycosylation pattern of individual proteins, exhibiting larger variability in disease states, have been suggested as biomarkers for different types of cancer, as well as inflammatory and neurodegenerative diseases. In this paper, we present a rapid and efficient method for glycosylation analysis of individual proteins focusing on changes in the degree of fucosylation or other alterations to the core structure of the glycans, such as the presence of bisecting N-acetylglucosamines and a modified degree of branching. Streptavidin-coated magnetic beads are used in combination with genetically engineered immunoaffinity binders, called VHH antibody fragments. A major advantage of the VHHs is that they are nonglycosylated; thus, enzymatic release of glycans from the targeted protein can be performed directly on the beads. After deglycosylation, the glycans are analyzed by MALDI-TOF-MS. The developed method was evaluated concerning its specificity, and thereafter implemented for studying the glycosylation pattern of two different proteins, alpha-1-antitrypsin and transferrin, in human serum and cerebrospinal fluid. To our knowledge, this is the first example of a protein array-type experiment that employs bead-based immunoaffinity purification in combination with mass spectrometry analysis for fast and efficient glycan analysis of individual proteins in biological fluid., (Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.)

Fatigue is a very common non-motor symptom in Parkinson disease (PD) patients. It included physical fatigue and mental fatigue. The potential mechanisms of mental fatigue involving serotonergic dysfunction and abnormal iron metabolism are still unknown. Therefore, we evaluated the fatigue symptoms, classified PD patients into fatigue group and non-fatigue group, and detected the levels of serotonin, iron and related proteins in CSF and serum. In CSF, 5-HT level is significantly decreased and the levels of iron and transferrin are dramatically increased in fatigue group. In fatigue group, mental fatigue score is negatively correlated with 5-HT level in CSF, and positively correlated with the scores of depression and excessive daytime sleepiness, and disease duration, also, mental fatigue is positively correlated with the levels of iron and transferrin in CSF. Transferrin level is negatively correlated with 5-HT level in CSF. In serum, the levels of 5-HT and transferrin are markedly decreased in fatigue group; mental fatigue score exhibits a negative correlation with 5-HT level. Thus serotonin dysfunction in both central and peripheral systems may be correlated with mental fatigue through abnormal iron metabolism. Depression, excessive daytime sleepiness and disease duration were the risk factors for mental fatigue of PD.

Two transferrin (Tf) glycan-isoforms were previously found in cerebrospinal fluid (CSF); one appears to be derived from serum (Tf-2) and the other from choroid plexus, a CSF-producing tissue (Tf-1). To analyse metabolic differences associated with the two isoforms, their ratio (Tf-2/Tf-1) was defined as the Tf index. Here we report that Tf indices of patients with tauopathies including Alzheimer's disease (2.29 + 0.64) were similar to those of neurological controls (2.07 + 0.87) (P = 0.147). In contrast, Tf indices with Parkinson's disease (PD, 3.38 ± 1.87) and multiple system atrophy (MSA, 3.15 ± 1.72) were higher than those of the controls (2.07 ± 0.87), the P-values being < 0.001 and 0.024, respectively. Tf indices of PD and MSA did not appear to be normally distributed. Indeed, detrended normal Quantile-Quantile plot analysis revealed the presence of an independent subgroup showing higher Tf indices in PD and MSA. The subgroup of PD showed higher levels of CSF α-synuclein (38.3 ± 17.8 ng/ml) than the rest (25.3 ± 11.3 ng/ml, P = 0.012). These results suggest that PD (and MSA) includes two subgroups, which show different metabolism of CSF transferrin and α-synuclein., (© The Authors 2016. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.)

Objective: Cerebrospinal fluid (CSF) leaks may present as rhinorrhea or otorrhea and can lead to life-threatening complications if not detected. The usefulness of the morpholinopropanesulfonic acid (MOPS)-histidine buffer in detecting β₂-transferrin, which is only found in the cerebrospinal fluid, was compared with the standard barbital buffer., Methods: We evaluated 20 aural or nasal patient specimens submitted for CSF testing via agarose electrophoresis with barbital and MOPS-histidine buffers., Results: The MOPS-histidine and barbital buffers revealed 5 transferrin bands and 2 transferrin bands with CSF, respectively. Seventeen of 20 patient specimens had concordant results. The 3 discrepant specimens initially tested negative with the barbital buffer and positive with the MOPS-histidine buffer. Two of the 3 patient specimens later tested positive with the barbital buffer when a new specimen was submitted., Conclusion: Agarose electrophoresis with the MOPS-histidine buffer increases the resolution of transferrin isoforms., (Copyright© by the American Society for Clinical Pathology (ASCP).)

A simple and rapid detection of cerebrospinal fluid (CSF) leakage would benefit spine surgeons making critical postoperative decisions on patient care. We have assessed novel approaches to selectively determine CSF β2-transferrin (β2TF), an asialo-transferrin (aTF) biomarker, without interference from serum sialo-transferrin (sTF) in test samples. First, we performed mild periodate oxidation to selectively generate aldehyde groups in sTF for capture with magnetic hydrazide microparticles, and selective removal with a magnetic separator. Using this protocol sTF was selectively removed from mixtures of CSF and serum containing CSF aTF (β2TF) and serum sTF, respectively. Second, a two-step enzymatic method was developed with neuraminidase and galactose oxidase for generating aldehyde groups in sTF present in CSF and serum mixtures for magnetic hydrazide microparticle capture. After selectively removing sTF from mixtures of CSF and serum, ELISA could detect significant TF signal only in CSF, while the TF signal in serum was negligible. The new approach for selective removal of only sTF in test samples will be promising for the required intervention by a spine surgeon., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Objectives/hypothesis: The objectives of this study were to examine the presence of β-2 transferrin (β2TRNSF) in cerebrospinal fluid (CSF) contaminated in vitro by various bacteria and explore the mechanism (passive or active) responsible for β2TRNSF elimination. Early diagnosis of CSF leakage may change treatment decisions and minimize the risk of meningitis and encephalitis. β2TRNSF 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 ability to cause neurosurgical-related infections: Streptococcus pneumoniae, methicillin-sensitive Staphylococcus aureus, Staphylococcus epidermidis, and Pseudomonas aeruginosa. β2TRNSF was examined at two time points: following immediate inoculation (t0) and following an overnight incubation (t18) over various bacterial concentrations. Samples of CSF inoculated with S pneumoniae were also examined in the presence of ciprofloxacin. For β2TRNSF analysis we used immunoblotting electrophoresis and enzyme-linked immunosorbent assay (ELISA)., Results: CSF samples collected from nine patients were analyzed. β2TRNSF was not detected following S pneumoniae inoculation at both time points when immunoblotting electrophoresis was used. Quantitative analysis using ELISA demonstrated significant β2TRNSF concentration decrease. The addition of ciprofloxacin led to the same results., Conclusions: CSF leak detection using β2TRNSF may be deceiving in the presence of a S pneumoniae cerebral nervous system infection. A passive process is suggested, as β2TRNSF disappeared either immediately or following incubation with inactive bacteria., (© 2014 The American Laryngological, Rhinological and Otological Society, Inc.)

Transferrin (Tf) is an 80 kDa iron-binding protein that is viewed as a promising drug carrier to target the central nervous system as a result of its ability to penetrate the blood-brain barrier. Among the many challenges during the development of Tf-based therapeutics, the sensitive and accurate quantitation of the administered Tf in cerebrospinal fluid (CSF) remains particularly difficult because of the presence of abundant endogenous Tf. Herein, we describe the development of a new liquid chromatography-mass spectrometry-based method for the sensitive and accurate quantitation of exogenous recombinant human Tf in rat CSF. By taking advantage of a His-tag present in recombinant Tf and applying Ni affinity purification, the exogenous human serum Tf can be greatly enriched from rat CSF, despite the presence of the abundant endogenous protein. Additionally, we applied a newly developed (18)O-labeling technique that can generate internal standards at the protein level, which greatly improved the accuracy and robustness of quantitation. The developed method was investigated for linearity, accuracy, precision, and lower limit of quantitation, all of which met the commonly accepted criteria for bioanalytical method validation.

Aims: Most biomarkers used for the premortem diagnosis of sporadic Creutzfeldt-Jakob disease (CJD) are surrogate in nature, and provide suboptimal sensitivity and specificity., Results: We report that CJD-associated brain iron dyshomeostasis is reflected in the cerebrospinal fluid (CSF), providing disease-specific diagnostic biomarkers. Analysis of 290 premortem CSF samples from confirmed cases of CJD, Alzheimer's disease, and other dementias (DMs), and 52 non-DM (ND) controls revealed a significant difference in ferroxidase (Frx) activity and transferrin (Tf) levels in sporadic Creutzfeldt-Jakob disease (sCJD) relative to other DM and ND controls. A combination of CSF Frx and Tf discriminated sCJD from other DMs with a sensitivity of 86.8%, specificity of 92.5%, accuracy of 88.9%, and area-under-the receiver-operating-characteristic (ROC) curve of 0.94. This combination provided a similar diagnostic accuracy in discriminating CJD from rapidly progressing cases who died within 6 months of sample collection. Surprisingly, ceruloplasmin and amyloid precursor protein, the major brain Frxs, displayed minimal activity in the CSF. Most of the Frx activity was concentrated in the <3-kDa fraction in normal and diseased CSF, and resisted heat and proteinase-K treatment., Innovation: (i) A combination of CSF Frx and Tf provides disease-specific premortem diagnostic biomarkers for sCJD. (ii) A novel, nonenzymatic, nonprotein Frx predominates in human CSF that is distinct from the currently known CSF Frxs., Conclusion: The underlying cause of iron imbalance is distinct in sCJD relative to other DMs associated with the brain iron imbalance. Thus, change in the CSF levels of iron-management proteins can provide disease-specific biomarkers and insight into the cause of iron imbalance in neurodegenerative conditions.

Objective: The aim of this study was to evaluate the CEofix™ CDT Capillary Electrophoresis (CE) kit for the detection of β-2-tranferrin (β-2-Tf) in cerebrospinal fluid (CSF)., Design and Method: Evaluation was performed according to CLSI EP5-A and EP12-A guidelines., Results: The method resolved β-2-Tf from other Tf isoforms. The C50 for β-Tf was determined to be 0.46 mg/L. Neither hemoglobin nor bilirubin co-migrated with β-2-Tf., Conclusions: The CDT kit can be used for detecting β-2-Tf in CSF by CE., (© 2013.)

We developed a high-throughput Enzyme-linked immunosorbent assay (ELISA) for measuring α2,6-sialylated transferrin (Tf), based on inhibition of anti-Tf antibody binding to α2,6-sialylated Tf by a lectin, Sambucus sieboldiana Agglutinin (SSA). The inhibition was not observed with other glycoforms, such as periodate-treated, sialidase-treated and sialidase/galactosidase-treated Tf, suggesting that the assay was glycoform specific. This finding was applied to an automated latex-agglutination immunoassay, using SSA lectin as an inhibitor (SSA-ALI). The concentration of α2,6-sialylated Tf measured by SSA-ALI in human cerebrospinal fluid was correlated with that of ELISA (r2 = 0.8554), previously developed for measuring α2,6-sialylated Tf.

Spontaneous rhinoliquorrhea with or without meningo-encephaloceles in the region of the sphenoid sinus occurs very infrequently. It is not uncommon that the attempt of transnasal endoscopic duraplasty in this region leads to recurrence of the CSF leak. The existence of a lateral craniopharyngeal canal can be a possible explanation for these failures.Retrospective analysis of 23 patients with rhinoliquorrhea of different pathogenesis in the region of the frontal and central skull base that were treated with transnasal, video-endoscopic surgical procedures in our department between 2006 and 2011.2 of 23 patients with proven rhinoliquorrhea following a transnasal video endoscopic duraplasty procedure showed a recurrence of the CSF leak. The computertomographic analysis with respect to the current literature indicated the presence of a craniopharyngeal canal at the lateral side of the sphenoid sinus. This canal is also known in the literature as Sternberg's canal. In contrast to the other 21 treated cases there were no planar skull base defects of different pathogenesis in these 2 cases, but a ontogenetically bony canal. The canal can reopen spontaneously or due to an external mechanical impact.The closure of this bony canal requires a modified surgical procedure such as sufficient padding of the bony canal and its sealing by a vascularized pedicle flap in contrast to the ordinary planar bony skull base defects., (© Georg Thieme Verlag KG Stuttgart · New York.)

A 73-year-old woman underwent routine enucleation for a blind, painful eye related to end-stage diabetic retinopathy and neovascular glaucoma. A large cystoid space, in continuity with the optic nerve stump, formed around the implant in the first few weeks following surgery. Aspirated contents were positive for β-2 transferrin, confirming cerebrospinal fluid origin. Multiple comorbidities delayed surgical intervention, but the condition was ultimately managed with exposure of the patent optic nerve sheath at the compartment's base, temporary control of cerebrospinal fluid leakage with pulmonary hyperventilation and topical fibrin glue, dissection and vascular-clip ligation of the nerve stump, and capping with a dermis-fat graft. To the authors' knowledge, this postenucleation entity has not been previously described, and asymptomatic idiopathic intracranial hypertension may have been an underlying factor.

Idiopathic normal pressure hydrocephalus (iNPH) is an elderly dementia caused by abnormal metabolism in the cerebrospinal fluid (CSF). The tap test is the current basis for confirming iNPH, but it shows very low sensitivity, indicating that many patients who might be cured by a shunt operation will be missed. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we found two transferrin isoforms: one had a unique N-glycan (Tf-1) whereas the other had N-glycan similar to that of serum transferrin (Tf-2). Glycan analyses revealed that Tf-1 had branching (biantennary) asialo- and agalacto-complex type N-glycans (N-acetylglucosamine [GlcNAc]-terminated glycans), which carried bisecting β1,4-N-acetylglucosamine and core α1,6-fucose. To examine glycoform whether changes in iNPH, we introduced the Tf-2/Tf-1 ratio as a diagnostic index, which minimized blot-to-blot variations in measurement. The Tf-2/Tf-1 ratios of iNPH patients are significantly higher than those of controls (p = 0.0019) and Alzheimer's patients (p = 0.0010). This suggests that the Tf-2/Tf-1 ratio could distinguish iNPH from Alzheimer's disease, and possibly other dementias. In conclusion, glycoform analysis has diagnostic potential in neurological diseases., (Copyright © 2012. Published by Elsevier Inc.)

Background: Transferrin is synthesized in the brain by choroid plexus and oligodendrocytes, but only that in the choroid plexus is secreted. Transferrin is a major iron delivery protein to the brain, but the amount transcytosed across the brain microvasculature is minimal. Transferrin is the major source of iron delivery to neurons. It may deliver iron to immature oligodendrocytes but this trophic effect declines over time while iron requirements for maintaining myelination continue. Finally, transferrin may play an important role in neurodegenerative diseases through its ability to mobilize iron., Scope of Review: The role of transferrin in maintaining brain iron homeostasis and the mechanism by which it enters the brain and delivers iron will be discussed. Its relevance to neurological disorders will also be addressed., Major Conclusions: Transferrin is the major iron delivery protein for neurons and the microvasculature, but has a limited role for glial cells. The main source of transferrin in the brain is likely from the choroid plexus although the concentration of transferrin at any given time in the brain includes that synthesized in oligodendrocytes. Little is known about brain iron egress or the role of transferrin in this process., General Significance: Neuron survival requires iron, which is predominantly delivered by transferrin. The concentration of transferrin in the cerebrospinal fluid is reflective of brain iron availability and can function as a biomarker in disease. Accumulation of iron in the brain contributes to neurodegenerative processes, thus an understanding of the role that transferrin plays in regulating brain iron homeostasis is essential. This article is part of a Special Issue entitled Transferrins: Molecular mechanisms of iron transport and disorders., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A Japanese male developed deafness, pyramidal signs and ataxia at age 50. A cerebrospinal fluid examination showed elevated levels of iron, transferrin and ferritin. Brain MRI showed atrophy of the cerebellum and pons as well as potential iron deposits on the surface of the brain. At autopsy, the brain weighed 1090 g and showed severe atrophy and necrosis of the cerebellum. No vascular malformation was observed. Extensive deposits of hemosiderin that were well stained with Berlin blue and ferritin immunohistochemistry were present at the surface and in the superficial layers of the cerebrum, brainstem, cerebellum and spinal cord. In these regions, numerous AT8 (p-τ)-immunopositive deposits were present in neurons and glia. In addition, phosphorylated α-synuclein-immunopositive Lewy bodies and neurites were observed in the brainstem nuclei. In the present report, the authors derive the novel insight that superficial siderosis is a distinctive entity associated with tauopathy and synucleinopathy.

Sporadic Creutzfeldt-Jakob-disease (sCJD) is a fatal neurodegenerative condition that escapes detection until autopsy. Recently, brain iron dyshomeostasis accompanied by increased transferrin (Tf) was reported in sCJD cases. The consequence of this abnormality on cerebrospinal-fluid (CSF) levels of Tf is uncertain. We evaluated the accuracy of CSF Tf, a 'new' biomarker, as a pre-mortem diagnostic test for sCJD when used alone or in combination with the 'current' biomarker total-tau (T-tau). Levels of total-Tf (T-Tf), isoforms of Tf (Tf-1 and Tf-β2), and iron saturation of Tf were quantified in CSF collected 0.3-36 months before death (duration) from 99 autopsy confirmed sCJD (CJD+) and 75 confirmed cases of dementia of non-CJD origin (CJD-). Diagnostic accuracy was estimated by non-parametric tests, logistic regression, and receiver operating characteristic (ROC) analysis. Area under the ROC curve (AUC), sensitivity, specificity, positive and negative predictive values (PV), and likelihood ratios (LR) of each biomarker and biomarker combination were calculated. We report that relative to CJD-, CJD+ cases had lower median CSF T-Tf (125,7093 vs. 217,7893) and higher T-tau (11530 vs. 1266) values. AUC was 0.90 (95% confidence interval (CI), 0.85-0.94) for T-Tf, and 0.93 (95% CI, 0.89-0.97) for T-Tf combined with T-tau. With cut-offs defined to achieve a sensitivity of ∼85%, T-Tf identified CJD+ cases with a specificity of 71.6% (95% CI, 59.1-81.7), positive LR of 3.0 (95% CI, 2.1-4.5), negative LR of 0.2 (95% CI, 0.1-0.3), and accuracy of 80.1%. The effect of patient age and duration was insignificant. T-Tf combined with T-tau identified CJD+ with improved specificity of 87.5% (95%CI, 76.3-94.1), positive LR of 6.8 (95% CI, 3.5-13.1), negative LR of 0.2 (95% CI, 0.1-0.3), positive-PV of 91.0%, negative-PV of 80.0%, and accuracy of 86.2%. Thus, CSF T-Tf, a new biomarker, when combined with the current biomarker T-tau, is a reliable pre-mortem diagnostic test for sCJD.

Objective: Detection of beta-2 transferrin in rhinorrhea fluid is a sensitive and specific method for the diagnosis of a cerebrospinal fluid (CSF) leak. Patients may be asked to collect this fluid at home to obtain an adequate volume for detection, and thus the age and storage conditions of these specimens may be variable upon analysis. The purpose of this study is to understand how age, storage temperature, and exposure to mucus affect the ability to detect beta-2 transferrin in CSF., Study Design: Case series with planned data collection., Setting: Tertiary care university hospital., Subjects and Methods: This study consists of 6 patients undergoing endoscopic CSF leak repair. CSF was collected directly from a lumbar drain (n = 4) or from nasal drainage (n = 2). Specimens were stored at 4°C (n = 3) or room temperature (n = 3). Samples were tested for the presence of beta-2 transferrin for up to 7 days using standard immunofixation electrophoresis techniques., Results: Beta-2 transferrin was detected in all specimens through day 7 regardless of storage temperature or collection site (95% exact binomial confidence interval of 0%-46%)., Conclusions: Beta-2 transferrin remains detectable in extracorporeal CSF for up to 7 days regardless of storage at room temperature or exposure to nasal mucus. Negative detection in patient specimens up to a week old is therefore not likely to be caused by protein degradation.

Asialo-Transferrin (S(0)) or beta(2)-Transferrin is an accepted marker of cerebrospinal fluid (CSF) leakage from the subarachnoid space into the nasal or aural cavity as the result of a head trauma. At this time, the S(0) detection method of choice is isoelectric focusing on polyacrylamide gel with direct immunofixation of transferrin (Tf) and silverstaining which are time-consuming techniques (> 5 h). The aim of this study is to present an alternative methodology for determination of Tf sialoforms present in CSF samples, specifically S(0), based on the detection of the Fe associated with those forms. A double spiking methodology is developed by saturating the protein with isotopically enriched iron ((57)Fe) and conducting post-column Isotope Dilution Analysis (IDA) with (54)Fe after separation of the different sialoforms by anion exchange chromatography. The results obtained have been validated using a certified serum (ERM-DA470) and applied with satisfactory results to a pooled CSF sample.

Background: Identification of the content of asialotransferrin in the cerebrospinal fluid is a diagnostic method for childhood-onset ataxia and central nervous system hypomyelination (CACH), also known as vanishing white matter disease (VWM), and also for other types of CNS disorders., Methods: In our work, we have determined the value of the ratio of the asialo form of transferrin to the total transferrin in the CSF using the commercially used Variant(TM) Bio-Rad system for the determination of carbohydrate-deficient transferrin (CDT) in serum. The peak corresponding to the asialo form of transferrin was identified with electrophoresis with subsequent immunofixation and mass spectrometry (MALDI-TOF/TOF)., Results: The intra-assay and inter-assay variations of the asialotransferrin value in CSF were 6.8% and 10.2%, respectively. Analysing CSF samples of 60 subjects (23 men aged 22-68 years and 37 women aged 18-77 years) with normal transferrin values and normal cytology as well as biochemistry parameters in the cerebrospinal fluid, and without apparent signs of neurological disorders, we have found the presence of 25.2 +/- 8.2% asialotransferrin., Conclusion: Except for the need to obtain approximately 1.5 mL of cerebrospinal fluid and a tenfold concentrating of the sample, there is no need to conduct any modifications of the preparation procedure for the analytic sample and chromatographic separation normally used for serum samples. The HPLC method of asialotransferrin determination in CSF provides clinically useful results.

In order to identify new metabolic abnormalities in patients with complex neurodegenerative disorders of unknown aetiology, we performed high resolution in vitro proton nuclear magnetic resonance spectroscopy on patient cerebrospinal fluid (CSF) samples. We identified five adult patients, including two sisters, with significantly elevated free sialic acid in the CSF compared to both the cohort of patients with diseases of unknown aetiology (n = 144; P < 0.001) and a control group of patients with well-defined diseases (n = 91; P < 0.001). All five patients displayed cerebellar ataxia, with peripheral neuropathy and cognitive decline or noteworthy behavioural changes. Cerebral MRI showed mild to moderate cerebellar atrophy (5/5) as well as white matter abnormalities in the cerebellum including the peridentate region (4/5), and at the periventricular level (3/5). Two-dimensional gel analyses revealed significant hyposialylation of transferrin in CSF of all patients compared to age-matched controls (P < 0.001)--a finding not present in the CSF of patients with Salla disease, the most common free sialic acid storage disorder. Free sialic acid content was normal in patients' urine and cultured fibroblasts as were plasma glycosylation patterns of transferrin. Analysis of the ganglioside profile in peripheral nerve biopsies of two out of five patients was also normal. Sequencing of four candidate genes in the free sialic acid biosynthetic pathway did not reveal any mutation. We therefore identified a new free sialic acid syndrome in which cerebellar ataxia is the leading symptom. The term CAFSA is suggested (cerebellar ataxia with free sialic acid).

The use of ESI-Q-TOF in combination with theoretical mapping of possible glycan variants by Diophantine mass analysis allows identification of transferrin (Tf) glycoforms in different biological fluids including human serum and cerebrospinal fluid (CSF). In order to restrict the structural variations, a chromatographic separation (HPLC) of the forms with different number of sialic acids is initially conducted. The individual fractions are purified, preconcentrated, and analyzed by ESI-Q-TOF. The results obtained experimentally are compared with those predicted by Diophantine calculations and the structures proposed. Each one of the found masses can be ascribed to a single Tf glycoform with a mass difference that ranged between -1 and -26 Da (average masses). Thus, the methodology can be used for structural characterization of Tf glycoforms relevant for clinical diagnosis without enzymatic digestion.

Cerebrospinal fluid (CSF) rhinorrhea is an uncommon but important medical condition. It can result from trauma, intracranial hypertension, or be idiopathic in origin. If left untreated, significant sequelae can result including infectious meningitis. Beta-2 transferrin is a sensitive and specific protein marker for CSF. Various radiographic modalities have been used to localize defects, including computed tomography (CT), magnetic resonance imaging, and CT cisternography. In recent years, surgical management of this condition has evolved significantly, primarily being performed endoscopically. Reconstruction can be performed with fat, free mucosal, or fascial grafts, or with vascularized flaps. The endoscopic surgeon should have a thorough understanding of the pathophysiology and diagnosis of CSF rhinorrhea as well as several surgical options in his or her armamentarium available to treat the patient suffering from CSF rhinorrhea.

So far only the detection of 14-3-3 proteins in cerebrospinal fluid (CSF) is included in the diagnostic criteria for sporadic Creutzfeldt-Jakob disease (sCJD). However, this assay cannot be used for screening because of the high rate of false positive results in sCJD, and often negative results in variant CJD. To facilitate the differential diagnosis of CJD, we applied 2-D differential gel-electrophoresis (2-D DIGE) as a quantitative proteomic screening system for CSF proteins. We compared 36 patients suffering from sCJD with 30 patients suffering from other neurodegenerative diseases. Sample preparation was optimized in consideration of the fact that CSF is composed of blood- and brain-derived proteins, and an improved 2-D DIGE protocol was established. Using this method in combination with protein identification by MALDI-TOF-MS, several known surrogate markers of sCJD like 14-3-3 protein, neuron-specific enolase, and lactate dehydrogenase were readily identified. Moreover, a not yet identified protein with an approximate molecular mass of 85 kDa was found as marker for sCJD with high diagnostic specificity and sensitivity. We conclude that our proteomic approach is useful to differentiate CJD from other neurodegenerative diseases and expect that CSF-optimized 2-D DIGE will find broad application in the search for other brain derived proteins in CSF.

Objective: This is a study estimating diagnostic accuracy of CSF asialotransferrin to transferrin ratio measurement in eIF2B related disorders by using clinical evaluation and EIF2B mutation analysis as the reference standard. eIF2B-related disorder is a relatively common leukodystrophy with broad phenotypic variation that is caused by mutations in any of the five EIF2B genes. There is a need for a simple and clinically valid screening tool for physicians evaluating patients with an unclassified leukodystrophy., Methods: CSF two-dimensional gel (2DG) electrophoresis analyses to measure asialotransferrin to transferrin ratios were performed in 60 subjects including 6 patients with documented EIF2B gene mutations, patients with other types of leukodystrophy, and patients with no leukodystrophy., Results: All six patients with mutation proven eIF2B-related disease showed low to nearly undetectable amounts of asialotransferrin in their CSF when compared to 54 unaffected controls by CSF 2DG analyses in this study. eIF2B-like patients, with clinically similar presentations but no mutations in EIF2B1-5, were distinguished from patients with mutations in EIF2B1-5 by this biomarker. Patients with mutations in EIF2B1-5 had asialotransferrin/transferrin ratio levels significantly different from the group as a whole (p < 0.001). Using 8% asialotransferrin/transferrin ratio as a cutoff, this biomarker has a 100% sensitivity (95% CI = 52-100%) and 94% specificity (95% CI = 84-99%)., Conclusion: Decreased asialotransferrin/transferrin ratio in the CSF of patients with eIF2B-related disorder is highly sensitive and specific. This rapid (<48 hours) and inexpensive diagnostic tool for eIF2B-related disorders has the potential to identify patients with likely eIF2B-related disorder for mutation analysis.

Background/aims: Alzheimer's disease (AD) is a well-known type of dementia. However, it remains difficult to identify AD in the early stage and to distinguish it from other dementing disorders. We examined glycoproteins in cerebrospinal fluid (CSF) as potential biological markers of AD., Methods: CSF samples were collected from AD, other dementia and nondemented patients. Glycoproteins in CSF were detected by lectin blotting using wheat germ agglutinin (WGA), and sugar chain analysis was performed by isoelectric focusing., Results: In Alzheimer's CSF, several glycoproteins had lower WGA-binding activities, one of which was sufficiently sensitive and specific to distinguish AD from nondemented controls and other dementias. Further analysis identified this glycosylated protein as transferrin, and altered sugar chain composition of transferrin isoforms was observed despite normal protein levels in CSF., Conclusion: The decreased WGA-binding activity of transferrin in AD is probably due to altered glycosylation of transferrin molecules. Transferrin glycosylation is thus a potential biological marker for AD diagnosis, and changes in this glycosylation may play an important role in the pathophysiology of AD., (Copyright 2008 S. Karger AG, Basel.)