Intrathecal synthesis of free immunoglobulin light chains in multiple sclerosis

Objective – The detection of oligoclonal immunoglobulin free light chains (FLC) in the diagnosis of multiple sclerosis (MS) was compared to IgG isoelectric focusing. Material and methods – Cerebrospinal fluid and serum samples from 69 patients with possible first attacks of MS, 50 patients with clinically definite MS (CDMS), and 118 patients with other neurological diseases (OND) were analyzed. IgG and FLC oligoclonal bands were detected by isoelectric focusing and immunoperoxidase staining. Results – Intrathecal synthesis of IgG, kappa FLC, and lambda FLC oligoclonal bands, respectively, was seen in 92%, 92%, and 86% of MS patients; in 61%, 62%, and 64% of patients with possible first attacks of MS; and in 3%, 3%, and 8% of the patients with OND. In control patients without IgG synthesis intrathecal lambda FLC synthesis was more common than kappa FLC synthesis ( P =0.03). Conclusion – Kappa FLC detection proved as useful as IgG analysis for the laboratory diagnosis of MS whereas the presence of intrathecal lambda FLC synthesis was less specific.     

寡克隆带和IgG鞘内合成率对多发性硬化的诊断价值

目的探讨寡克隆带(OCBs)和IgG鞘内合成率(IgGSyn)对多发性硬化(MS)诊断的敏感性、特异性,以及定性和定量指标的相关性。方法选取30例MS(MS组)、40例神经系统炎性疾病(NID组)和22例神经系统非炎性疾病(NNID组)患者,应用速率散射比浊法测定血清和脑脊液(CSF)中免疫球蛋白G(IgG)、白蛋白(Alb)水平,等电聚焦结合银染色法检测CSF中OCBs,计算IgGSyn,并对其敏感性、特异性和阳性结果似然比(PRLR)进行分析。结果OCBs阳性率和IgGSyn异常率MS组与NID组比较差异无显著性;MS组、NID组与NNID组比较差异有极显著性(均P<0.01)。MS组和NID组中,OCBs阳性者与阴性者IgGSyn值差异无显著性。对MS诊断的敏感性、特异性和PRLR,OCBs分别为63.3%、77.7%和2.8;IgGSyn为46.7%、75.2%和1.9。结论OCBs和IgGSyn检测结果的不完全一致性提示中枢神经系统内存在不同的体液免疫反应机制,综合分析OCBs和IgGSyn,对MS诊断具有参考价值。     

Detection of oligoclonal IgG bands in unconcentrated CSF by isoelectric focusing in agarose gel and silver staining

We analyzed 102 unconcentrated cerebrospinal fluids from a variety of neurologic diseases for oligoclonal IgG bands by isoelectric focusing in agarose gel followed by staining with silver. Ten to 12 microliter of cerebrospinal fluid containing 0.4-0.8 microgram of IgG was found to be optimum. Cerebrospinal fluid from 38 of 40 patients with clinically definite multiple sclerosis, 6 of 9 with suspected multiple sclerosis and 8 of 53 patients with other neurologic diseases had oligoclonal IgG bands by IEF in agarose followed by immunofixation. The commercial system employed here is a simple sensitive and rapid method for detection of oligoclonal bands in unconcentrated cerebrospinal fluid of patients with multiple sclerosis.     

Intrathecal IgG synthesis and autoantibody-secreting cells in multiple sclerosis

We studied intrathecal IgG synthesis and autoantibody-secreting cells in 148 patients with possible onset symptoms of MS (POSMS) or clinically definite MS (CDMS). In POSMS intrathecal synthesis of IgG oligoclonal bands and abnormalities on T2-weighted magnetic resonance imaging were associated but the former were more prevalent. The cerebrospinal fluid (CSF) leukocyte count and the number of anti-protelipid protein antibody-secreting cells in cerebrospinal fluid (CSF) correlated with disease activity in POSMS. Intrathecal IgG synthesis levels and the number of anti-myelin basic protein antibody-secreting cells in CSF correlated with disease activity in CDMS. Our results support recent reports of pathogenetic heterogeneity and a pathogenetic role of the antibody response in MS.     

IgG synthesis within the central nervous system. Comparison of three formulas

IgG synthesis within the central nervous system is the expression of a local pathologic immune reaction. Its detection is of great importance in a large number of neurologic diseases, especially multiple sclerosis (MS). The results obtained from the study of 115 neurologic patients (52 patients with MS and 63 patients with other neurologic diseases) using three different mathematic formulas (Tourtellotte's formula, the index of Tibbling and coworkers, and Schuller and Sagar's formula) are presented and statistically analyzed. Despite their obvious differences, the results from these three formulas are in accordance in 76% of the patients observed. The clinical correlations of these formulas are discussed; none of them allows a specific diagnosis of MS. The formula of Schuller and Sagar seems the most sensitive, detecting local central nervous system IgG synthesis in 79% of patients with MS, compared with 54% of patients with MS utilizing the index of Tibbling and coworkers and 50% of patients with MS utilizing Tourtellotte's formula. No correlation was established between serum and cerebrospinal fluid albumin concentrations, nor between serum albumin and IgG levels in cerebrospinal fluid. Thus, no evidence exists to support the inclusion of variations in serum albumin levels in the calculation of intrathecal IgG synthesis. Furthermore, Schuller and Sagar's formula allows classification of neuroimmunologic diseases and the evaluation of antibody-specific activity (ASA) in locally produced IgG. The comparison of intrathecal IgG ASA with serum IgG ASA, which reflects local and general immunity, respectively, seems potentially important.     

寡克隆区带和IgG指数在多发性硬化中的诊断意义

目的研究寡克隆区带（OCBs）和IgG指数（IgGI）对多发性硬化（MS）诊断的敏感性及其影响因素。方法用等电聚焦结合银染色法检测30例MS、40例神经系统炎性疾病（NID）和22例神经系统非炎性疾病（NNID）患者CSF中OCBs，并计算IgG I。结果MS组和NID组比较OCBs阳性率、IgG I异常率均无显著性差异（P〉0．05）；MS组、NID组与NNID组比较。差异均有显著性（P〈0．05）；传统型MS和脊髓型MS比较，差异均无显著性（P〉0．05）。OCBs对MS诊断的敏感性、特异性和阳性结果似然比分别为63．3％、77．7％和2．8；IgG I分别为40．0％、76．7％和1．7。结论本地区MSOCBs阳性率和IgG I异常率较低，可能与遗传背景、疾病类型和药物应用有关，OCBs和IgG I对MS诊断具有相对特异性。     

Cerebrospinal fluid IgG profiles and oligoclonal bands in Chinese patients with multiple sclerosis

OBJECTIVES: To investigate the significance of oligoclonal bands (OCBs) and intrathecal IgG fractions (IgGIF) for the diagnosis of multiple sclerosis (MS) in northern China. MATERIALS AND METHODS: OCBs in cerebrospinal fluid from 30 patients with MS, 34 with other inflammatory neurological diseases (IND) and 22 with non-inflammatory neurological diseases (NIND) were detected using isoelectric focusing. IgGIF was calculated based on corresponding formula. RESULTS: There was no significant difference in the frequencies of positive OCBs and elevated IgGIF between the MS group and the IND group. Compared with NIND, the MS and IND groups had a significantly higher incidence of OCBs and elevated IgGIF. The sensitivity, specificity and positive result likelihood ratio of OCBs for the diagnosis of MS were 63.3%, 74.2% and 2.5 respectively; those of IgGIF were 36.7%, 84.5% and 2.4. CONCLUSIONS: The two parameters, OCBs and IgGIF are of less diagnostic value for MS in China.     

Oligoclonal IgG bands in cerebrospinal fluid. Principles for demonstration and interpretation based on findings in 1114 neurological patients

Unconcentrated cerebrospinal fluid (CSF) and serum samples from 1114 consecutive patients were examined for presence of oligoclonal IgG bands (OB) by agarose isoelectric focusing (AIF) followed by protein transfer to nitrocellulose membrane, immunolabeling, and avidinbiotin-peroxidase staining (avidin-biotin AIF). Oligoclonal bands were demonstrated in CSF from all 58 patients with multiple sclerosis (MS), eight of 29 with aseptic nervous system infections, and 9% of 1014 with other neurological disorders (OND) considered as noninflammatory at primary clinical evaluation. Comparative examination of all specimens in another laboratory by conventional AIF after concentration of CSF revealed lower frequencies of OB in all diagnostic groups. In addition to the high sensitivity of avidinbiotin AIF, which enables detection of OB by separation of 5 microL of unconcentrated CSF even when the CSF IgG level is around the lower normal range, the procedure also has optimal specificity since IgG exclusively is detected. Avidin-biotin AIF may be the method preferred for routine examination of CSF for OB. Demonstration of OB in CSF is valuable especially in MS, where, in contrast to diagnostic aids such as evoked potentials and neuro-imaging, it establishes inflammatory type of nervous system involvements. Oligoclonal IgG bands in CSF from patients with OND reflect intrathecal immune response and should lead to investigations of infectious etiology.     

CSF寡克隆区带对多发性硬化的诊断意义

目的:琼脂糖等电聚焦结合双抗体过氧化物酶标记、亲和素-生物素放大技术检测CSF寡克隆区带,研究该方法的敏感性和特异性。方法:琼脂糖等电聚焦结合双抗体过氧化物酶标记、亲和素-生物素放大技术检测21例MS患者、42例神经系统炎性疾病(NID)患者和19例神经系统非炎性疾病(NNID)患者的CSF及对照血清。结果:MS患者寡克隆区带阳性率为47.6%,诊断特异性为98.4%,MS组与NID组、NNID组的差异均有统计学意义(P<0.0125)。结论:该方法敏感、特异,是MS临床诊断最有价值的免疫学指标。我国MS患者寡克隆区带的阳性率较欧美地区低,但与亚洲一些国家及我国的台湾和香港地区接近,阳性率差异可能与东西方MS患者的免疫遗传背景不同有关。     

Intrathecal levels of vitamin D and IgG in multiple sclerosis

Holmøy T, Lossius A, Gundersen TE, Moen SM, Castellazzi M, Fainardi E, Casetta I. Intrathecal levels of vitamin D and IgG in multiple sclerosis.
Acta Neurol Scand: 2012: 125: e28–e31.
© 2011 John Wiley & Sons A/S. Background – Intrathecal synthesis of IgG is a hallmark of multiple sclerosis (MS). Vitamin D may modulate B‐cell function and dampen the synthesis of IgG. Objective – To investigate the relation between vitamin D levels in cerebrospinal fluid and serum and intrathecal synthesis of IgG. Methods – 25‐hydroxyvitamin D (25(OH)D) and IgG were assessed in cerebrospinal fluid and serum in 40 patients with MS. Results – There was no significant correlation between the IgG index and 25(OH)D levels in cerebrospinal fluid or serum. The levels of 25(OH)D in cerebrospinal fluid and serum did not differ between patients with and without intrathecal synthesis of IgG. There was a non‐significant trend towards a positive correlation between the concentrations of 25(OH)D and IgG in the cerebrospinal fluid, but not in serum. Conclusion – Physiological variation in vitamin D does not exert a major impact on intrathecal synthesis of IgG in MS.     

Multiple sclerosis. Cerebrospinal fluid immune complexes that bind C1q

We used a sensitive C1q-binding assay to measure levels of soluble immune complexes in 182 samples of cerebrospinal fluid (CSF) from control patients and patients with multiple sclerosis (MS). Soluble immune complexes in CSF were detected in 16% of patients with progressive MS, 38% of patients with exacerbating-remitting MS, 55% of patients with infectious or inflammatory diseases, 3% of patients with noninflammatory neurologic disorders, and in 0% of control patients with back pain. No correlations were found between the results of the C1q-binding assay and abnormalities of other CSF parameters. These included an elevated level of myelin basic protein, pleocytosis, oligoclonal bands, or an increased IgG level. Because of the lack of correlation to laboratory indexes of disease activity and the nonspecificity of a positive test, the C1q-binding assay seems to have little clinical usefulness in the diagnosis or management of patients with MS.     

Oligoclonal IgA bands in multiple sclerosis and subacute sclerosing panencephalitis

CSF oligoclonal IgG bands are often found in MS or cerebral diseases in which there is chronic antigenic stimulation. Using agarose isoelectric focusing followed by immunoblotting, we found oligoclonal IgA bands in CSF from 16 of 20 randomly selected patients with MS, 7 of 7 with subacute sclerosing panencephalitis (SSPE), and 0 of 10 with noninflammatory neurologic or psychiatric disease. IgA bands did not correlate with the course or stage of MS. Serial samples from two patients with MS and one with SSPE demonstrated only minor changes in IgA banding pattern. One MS patient without oligoclonal IgG bands had oligoclonal IgA bands, indicating that the latter test may be helpful in the diagnosis of MS.     

Analysis of Chlamydia pneumoniae-specific oligoclonal bands in multiple sclerosis and other neurologic diseases

OBJECTIVE: Paired serum and cerebrospinal fluid (CSF) specimens were investigated for Chlamydia pneumoniae-specific oligoclonal bands (OCBs) to determine band specificity in multiple sclerosis (MS). MATERIAL AND METHODS: Serum and CSF samples were collected from patients with relapsing-remitting MS (n = 56), other inflammatory (n = 18) or non-inflammatory (n = 15) neurologic diseases, and from 10 healthy controls. OCBs were determined with affinity immunoblotting of C. pneumoniae-specific IgG onto antigen-coated nitrocellulose paper after protein separation with agarose isoelectric focusing. RESULTS: Chlamydia pneumoniae-specific OCBs were present in 5 of 56 patients with MS, and in 3 of 18 patients with other inflammatory neurologic diseases. CONCLUSIONS: The intrathecal production of C. pneumoniae-specific oligoclonal IgG occurs in a minority of patients with MS. This intrathecal anti-C. pneumoniae reactivity is likely part of a polyspecific humoral immune response in MS.     

Increased peripheral blood CD5+ B cells predict earlier conversion to MS in high-risk clinically isolated syndromes

Clinically isolated syndrome patients (CIS) with oligoclonal IgG bands (OCGB) are at high risk for clinically definite multiple sclerosis (MS). However, the outcome for individual patients is unpredictable and the search for reliable blood markers predicting early conversion to multiple sclerosis (MS) has clinical relevance. CD5+ B cells (CD5+Bc) are involved in some autoimmune diseases. This study investigated whether high blood CD5+Bc percentage can predict CIS conversion to MS. Fifty-five consecutive CIS showing OCGB were prospectively studied. Every patient underwent a brain MRI study and a flow cytometry analysis of CD5+Bc percentage. Conversion to MS was studied during follow-up. The CD5+Bc percentage was assessed in 40 controls and a cut-off value of 3.5% (mean+2 SD) was calculated. A blood CD5+Bc percentage above this value predicted earlier conversion to MS in the whole group (hazard ratio [HR]: 3.40; 95% confidence interval [CI]: 1.69-6.68; p=0.0005) and in CIS patients fulfilling three or more Barkhof-Tintoré criteria plus OCGB, who showed higher risk for MS (HR: 3.79; 95% CI: 1.86-15.32; p=0.0018). Multivariate analysis also showed a predictive value for high blood CD5+Bc count (HR: 4.3; 95% CI: 1.9-9.5; p<0.0001). It was concluded that high percentages of CD5+Bc independently associate with increased risk of early conversion to MS in CIS patients with OCGB and Barkhof-Tintoré criteria.     

The cerebrospinal fluid in multiple sclerosis. Quantitative assessment of intrathecal immunoglobulin synthesis by empirical formulae

Intrathecal synthesis of IgG occurs in more than 90% of patients with clinically definite multiple sclerosis. The prevalence and significance of intrathecal synthesis of IgA and IgM are, however, less thoroughly characterized. We estimated intrathecal synthesis of IgG, IgA and IgM with various empirical formulae. The concentrations of albumin, IgG, IgA and IgM and the presence of IgG oligoclonal bands were determined in CSF and serum from 350 patients, including 97 with clinically definite multiple sclerosis. Intrathecal synthesis of IgG oligoclonal bands was detected in 95% of patients with multiple sclerosis (95% confidence interval 88–98%). The IgG-index, an extended IgG-index, and a hyperbolic IgG formula performed approximately equally in identifying patients with MS, but they were all inferior to the detection of IgG oligoclonal bands. In quantitative measurements, the extended immunoglobulin indices appeared to perform well; studies comparing the extended IgA- and IgM-indices to qualitative analyses (electrophoresis or isoelectric focusing) are, however, needed to confirm this. Detection of intrathecal synthesis of IgA of IgM was of little value in the diagnosis of multiple sclerosis.     

HHV‐6 infection in multiple sclerosis. A clinical and laboratory analysis

Background and purpose: To elucidate the role of human herpesvirus‐6 (HHV‐6) in the development of multiple sclerosis (MS). Patients and methods: Nine patients with MS and with acute or chronic HHV‐6 infection were evaluated. Results: Intrathecal antibody production to HHV‐6 and oligoclonal IgG bands in the cerebrospinal fluid (CSF) was observed in two patients with a clinically definite MS and chronic HHV‐6 infection (based on the presence of HHV‐6 specific antibodies in the CSF). A temporal association between the symptoms of clinically possible MS and acute primary HHV‐6A infection (based on avidity of HHV‐6 specific antibodies) was observed in two patients. Conclusions: Human herpesvirus‐6 infection may be an associated agent in some MS cases. Viral studies are needed to identify a possible viral etiology and give specific therapy.     

Isoelectric focusing is superior to immunofixation electrophoresis in diagnosing CNS inflammation

BACKGROUND: A sensitive method to detect intrathecal IgG production is important in diagnosing inflammatory central nervous system (CNS) diseases, including multiple sclerosis (MS). OBJECTIVE: To compare cerebrospinal fluid (CSF) electrophoresis with isoelectric focusing (IEF), immunofixation-peroxidase electrophoresis (IFPE) and high-resolution agarose electrophoresis with protein-staining (HRAGE). METHODS: Paired serum and CSF samples from 307 consecutive patients attending a general neurology clinic were examined with IEF, IFPE and HRAGE. Clinical diagnosis was based on review of the patients' medical records after an average of 4 years. RESULTS: The sensitivity for detecting any inflammatory (autoimmune or infectious) CNS disease (52 patients) was 67% for IEF, 50% for IFPE and 29% for HRAGE. The sensitivity for detecting MS (14 patients) was 93%, 86% and 29% respectively. The sensitivity for detecting clinically isolated syndrome (eight patients) was 75%, 25% and 13% respectively. The number of oligoclonal bands in IEF was higher in inflammatory than in non-inflammatory neurological diseases or symptoms, but similar in MS and other inflammatory diseases. CONCLUSION: IEF is the method of choice in diagnosing intrathecal IgG synthesis.     

Multiple sclerosis--studies of the correlations between selected parameters of immune response and the clinical course of the disease]

In the paper the following immune response parameters were analyzed: number of leucocytes in the cerebrospinal fluid, IgG index, presence and number of oligoclonal IgG bands. The correlation between these parameters and between the parameters and clinical course of multiple sclerosis was studied. Oligoclonal bands were present in the cerebrospinal fluid in 92% cases of MS, and that was the most frequent abnormality among immune response parameters studied. There was a correlation between IgG index, number of leucocytes in the cerebrospinal fluid and number of oligoclonal IgG bands in patients with relapsing-remitting course of the disease. We did not find any statistical differences between the number of leucocytes in the cerebrospinal fluid, the frequency of pleocytosis and the type of the disease, relapsing remitting or chronic progressive. The type of the disease, its duration and number of relapses did not significantly influence the IgG index values and number of IgG oligoclonal bands in the cerebrospinal fluid.     

Detection of oligoclonal free kappa chains in the absence of oligoclonal IgG in the CSF of patients with suspected multiple sclerosis

BACKGROUND: Oligoclonal free kappa bands are present as frequently as oligoclonal IgG bands in the cerebrospinal fluid (CSF) from patients with definite multiple sclerosis (MS) and can even occur in the absence of oligoclonal IgG. As such, they too are markers of an ongoing intrathecal immune process. OBJECTIVES: To determine how frequently oligoclonal free kappa bands are detectable in the CSF from patients with clinical signs and symptoms suggestive of MS in the absence of CSF restricted oligoclonal IgG. METHODS: An immunoaffinity mediated immunoblotting technique specific for free kappa chains was used, after isoelectric focusing of paired CSF and serum samples from 33 patients with clinical signs and symptoms suggestive of MS but without CSF oligoclonal IgG. CSF data were correlated with MRI results in the context of the new diagnostic criteria from McDonald et al. RESULTS: Eighteen CSF samples contained oligoclonal free kappa bands (54%), mainly from patients with motor dysfunction (83%) and optic neuritis (64%). All patients with a positive MRI according to Barkhof's criteria (n = 6) had free kappa bands in their CSF. CONCLUSIONS: (1) Oligoclonal free kappa bands in the CSF are related to the dissemination of MS lesions; (2) such bands should be looked for in oligoclonal IgG negative CSF, and (3) the presence of free kappa bands in the CSF may be a substitute for oligoclonal IgG in the McDonald's criteria for diagnosis of MS.     

Detection of oligoclonal IgG bands in unconcentrated CSF in multiple sclerosis and other neurological diseases by isoelectric focusing on ultrathin-layer polyacrylamide gel immunofixation and silver staining

Isoelectric focusing of proteins (IEF) in ultrathin-layer polyacrylamide gel (0.4 mm, PAG), followed by direct immunofixation with monospecific antiserum and silver staining, is a highly specific, sensitive and simple method for the demonstration of oligoclonal IgG in unconcentrated cerebrospinal fluid (CSF) samples (5-10 microliters). For the present method, the optimal concentrations of IgG in CSF samples are about 0.025-0.030 g/l, corresponding to the applied amount of 125-150 mg. In our testing of this method, oligoclonal IgG bands in CSF specimens were clearly demonstrated in 52 (96%) of 54 patients with clinically established definite diagnosis of multiple sclerosis (MS), in 4 (40%) of 10 patients with infectious diseases of the CNS, and in 9 patients (25%) of 38 with other neurological diseases. Abnormal patterns were also demonstrated in the serum of patients with MS (43%). Intrathecally synthesized IgG was mathematically calculated in 43 (80%) out of 54 patients with MS. This method appears to be a useful alternative for the demonstration of oligoclonal IgG bands in the unconcentrated CSF, especially when questionable or negative results arise by routine electrophoretic technique for oligoclonal bands detection.