Implications of monoclonal gammopathy and isoelectric focusing pattern 5 on the free light chain kappa diagnostics in cerebrospinal fluid.

OBJECTIVES: Oligoclonal bands (OCB) analysis is the reference standard for detecting an intrathecal IgG synthesis. Alongside OCB, free light chains kappa (FLCκ) are considered an additional sensitive biomarker for determining patterns 2 or 3, indicating intrathecal Ig synthesis. However, kFLC IF is not suitable for detecting a monoclonal pattern 5. The primary aim of this study was to evaluate the impact of incorporating FLCκ analysis into routine cerebrospinal fluid (CSF) diagnostics instead of OCB testing on the rate of missed monoclonal IgG detection. METHODS: A two-center retrospective biomarker study was conducted. OCB were identified using isoelectric focusing in polyacrylamide gels followed by silver staining or in agarose gels followed by immunofixation. FLCκ were quantified using nephelometry and FLCκ assay (Siemens). RESULTS: Out of a combined total of 17,755 OCB analyses conducted between 2011 and 2021, a subset of 269 cases (1.5 %) exhibited pattern 5. 98 samples (36 %), which included 18 samples with intrathecal inflammation as determined by additional OCB pattern 2 were included in the FLCκ analysis. Of those, 16 (89 %) had intrathecal FLCκ synthesis. CONCLUSIONS: While FLCκ offers a promising avenue for detecting an intrathecal inflammation, the pattern 5, though rare, remains a valuable additional finding of OCB analysis. A combined approach of FLCκ and OCB analysis is recommended for a comprehensive assessment of the humoral intrathecal immune response.

Interleukin-33 as a Biomarker Affecting Intrathecal Synthesis of Immunoglobulin in Neuromyelitis Optica Spectrum Disorder and Myelin Oligodendrocyte Glycoprotein Antibody-Associated Disease.

INTRODUCTION: The purpose of this study was to analyze IL-33 maybe as a biomarker especially with respect to intrathecal immunoglobulin G (IgG) synthesis which was involved in the immune-mediated process in the demyelinating disease of the central nervous system. METHODS: We aimed to determine the risk association of the serum and CSF levels of IL-33 in aquaporin-4 (AQP4)+neuromyelitis optica spectrum disorder (NMOSD) patients and myelin oligodendrocyte glycoprotein antibody-associated disease (MOGAD) patients compared with the control group. Levels of inflammatory (IL-2, IL-4, IL-6, and IL-10) markers and QAlb, the IgG index, and 24-h IgG synthesis rate were assessed in 28 AQP4+NMOSD patients and 11 MOGAD patients. Disease severity was assessed using the Expanded Disability Status Scale (EDSS). RESULTS: The level of IL-33 in serum decreased first but then increased gradually in AQP4+NMOSD and MOGAD. The serum level of IL-2, IL-4, and IL-10 increased more significantly and decreased more rapidly after methylprednisolone treatment. The level of IL-33 in CSF increased progressively in AQP4+NMOSD and MOGAD, especially in MOGAD. The QAlb levels were increased significantly in the CSF of MOGAD patients and AQP4+NMOSD patients on the acute stage of the disease. The IgG index and 24-h IgG synthesis rate were also increased significantly in the CSF of two groups similarly. CONCLUSIONS: Thus, we concluded that IL-33 may induce dysfunction of the blood-brain barrier and lead to intrathecal synthesis of immunoglobulin in the AQP4+NMOSD and MOGAD, especially in MOGAD. It maybe as a biomarker, at least in part, was involved in the demyelinating diseases of the central nervous system.

Spatial and temporal variation of routine parameters: pitfalls in the cerebrospinal fluid analysis in central nervous system infections.

The cerebrospinal fluid (CSF) space is convoluted. CSF flow oscillates with a net flow from the ventricles towards the cerebral and spinal subarachnoid space. This flow is influenced by heartbeats, breath, head or body movements as well as the activity of the ciliated epithelium of the plexus and ventricular ependyma. The shape of the CSF space and the CSF flow preclude rapid equilibration of cells, proteins and smaller compounds between the different parts of the compartment. In this review including reinterpretation of previously published data we illustrate, how anatomical and (patho)physiological conditions can influence routine CSF analysis. Equilibration of the components of the CSF depends on the size of the molecule or particle, e.g., lactate is distributed in the CSF more homogeneously than proteins or cells. The concentrations of blood-derived compounds usually increase from the ventricles to the lumbar CSF space, whereas the concentrations of brain-derived compounds usually decrease. Under special conditions, in particular when distribution is impaired, the rostro-caudal gradient of blood-derived compounds can be reversed. In the last century, several researchers attempted to define typical CSF findings for the diagnosis of several inflammatory diseases based on routine parameters. Because of the high spatial and temporal variations, findings considered typical of certain CNS diseases often are absent in parts of or even in the entire CSF compartment. In CNS infections, identification of the pathogen by culture, antigen detection or molecular methods is essential for diagnosis.

Diagnostic value of kappa free light chains in patients with one isolated band in isoelectric focusing.

OBJECTIVES: Oligoclonal band (OCB) determination in cerebrospinal fluid (CSF) is the gold standard to detect intrathecal inflammation. However, there is uncertainty about the significance of one isolated band in CSF. Free light chains kappa (FLC-k) are gaining interest as a complementary method to detect intrathecal inflammation. The aim of this study was to investigate the performance of an additive measurement of FLC-k in patients with one isolated band in CSF. MATERIALS & METHODS: FLC-k were analyzed using the nephelometric Siemens FLC-k kit in paired samples of CSF and sera (n = 56) in patients with one isolated band in isoelectric focusing. According to medical diagnosis, samples were subdivided in inflammatory neurological disease, non-inflammatory neurological disease controls and symptomatic controls. Intrathecal fraction of FLC-k was plotted in a FLC-k quotient diagram. OCB interpretation was done blinded by three experienced raters. RESULTS: Of 6695 OCB analyses, 91 (1.4%) had one isolated band in CSF. After exclusion of patient samples due to unclear OCB pattern after reevaluation and sample availability, 56 patient samples were included in the study. All patients with an inflammatory origin of disease (n = 13) had FLC-k values above the upper discrimination line (Qlim) in the FLC-k quotient diagram, resulting in a sensitivity of 100% with a positive predictive value of 52% and a negative predictive value of 100%. Fourteen patients (36%) with a non-inflammatory origin of disease (n = 39) had FLC-k values above Qlim. CONCLUSIONS: In patients with one isolated band in CSF, a lack of intrathecal fraction of FLC-k strongly favors a non-inflammatory orgin of disease. Implementation of FLC-k measurement can help the clinician in the diagnostic process of neurological diseases.

The Influence of Blood Contamination on Cerebrospinal Fluid Diagnostics.

Background: Blood contamination due to traumatic lumbar puncture presents a diagnostic pitfall in cerebrospinal fluid (CSF) analysis. It is controversially discussed if phagocytosis of erythrocytes which can be found in the CSF after subarachnoid hemorrhage can also develop in vitro in the presence of artificial blood contamination. Furthermore, there is no consensus about the acceptable amount of artificial blood contamination on CSF protein results. Methods: Two measurement series were performed in order to investigate the role of artificial blood contamination on the possible development of erythrophages and siderophages in the CSF: (1) blood contamination was simulated in vitro by adding blood into the CSF. (2) CSF was investigated when blood contamination occurred during a traumatic lumbar puncture. In both types of experiments, CSF including blood was incubated for 24 h and for 72 h at room temperature or at 4°C. In the third measurement series, the effects of artificial blood contamination on CSF protein results were investigated. Blood contamination was simulated in vitro by adding different amounts of blood ending up with five different samples containing erythrocyte counts of 2,500, 5,000, 7,500, 10,000, and 20,000 per µl CSF. Results: Cytological examination revealed no evidence of erythrophages or siderophages in vitro. In contrast, already a low blood contamination (2,500 erythrocytes/µl CSF) led to false pathological results of total protein and albumin. Along with increasing amounts of blood, the frequency of false pathological protein results increased. A blood contamination of 5,000 erythrocytes/µl CSF resulted in a false positive intrathecal IgM production in nearly every fifth patient. In contrast, blood contamination with 5,000 erythrocytes/µl CSF was the acceptable amount of blood which did not lead to a false positive intrathecal synthesis of IgG and IgA. Conclusion: Erythrophages and siderophages do not develop in vitro. An extensive diagnostic work up for the source of blood in the CSF should be performed when erythrophages or siderophages are found in the CSF. The contamination of CSF with increasing volume of blood resulted in falsely elevated CSF protein concentrations. Hence, the amount of blood contamination has to be taken into consideration when interpreting CSF protein measurement results.

Kappa free light chains in cerebrospinal fluid to identify patients with oligoclonal bands.

BACKGROUND AND PURPOSE: The gold standard for detection of intrathecal immunoglobulin synthesis is the measurement of oligoclonal bands (OCB). In the diagnosis of multiple sclerosis, the kappa free light chains (KFLC) index has a similar sensitivity and specificity as OCB. This study investigated whether determination of the KFLC index could be used to predict the presence of OCB. METHODS: The KFLC index was determined prospectively from 295 paired serum and cerebrospinal fluid samples. KFLC were determined by nephelometry using the N Latex FLC kappa kit (Siemens Healthcare Diagnostics Products GmbH) on the BN Prospec analyzer (Siemens Healthcare Diagnostics Products GmbH) (cohort I). A cut-off value was determined using receiver operating characteristic analysis in relation to OCB positivity. These results were validated prospectively in 96 samples (cohort II) as well as retrospectively in samples of 46 patients known to be OCB positive (cohort III). We also compared the agreement of two commercially available nephelometric KFLC assays. RESULTS: In cohort I, a KFLC index of 3.61 yielded 100% sensitivity and 88% specificity. Prospective validation of this cut-off value in cohort II showed 92% sensitivity and 96% specificity. In cohort III, a sensitivity of 93% was achieved. Comparison of Siemens and Binding Site (Birmingham, UK) assays revealed good agreement (r2  = 0.86). CONCLUSIONS: The KFLC index with a cut-off value of 3.61 had high diagnostic accuracy to predict immunoglobulin G synthesis via OCB analysis. Determination of the KFLC index provided a quantitative parameter that could be used as an initial diagnostic step in inflammatory central nervous system disorders before measuring OCB.