高频超声对上肢桡神经检测正常参考值的研究

目的测量正常人上肢桡神经在高频超声(HUS)下神经形态、声像学特点及横截面积(CSA)的正常值并探讨神经与周围组织的关系;为高频超声检侧外周神经疾病提供正常参考值。方法对临床检查正常的200例健康志愿者沿桡神经在神经预定的测量点上依次获取超声声像图及测量各点神经的横截面积,每个测量点重复测量三次取其平均值,并做神经CSA与身高、体重的相关性分析。结论高频超声下正常人上肢神经呈筛网状低回声图像,横截面呈圆形、卵圆形或三角形。测量桡神经各测量点处的神经横截面积,桡神经在肱骨外髁上4cm、桡神经沟处2点面积依次为(x珋±S)5.14±1.24 mm2、5.08±1.23mm2。左右上肢之间同一测量点比较神经横截面积P>0.05,桡神经的CSAs同年龄组男女之间比较P<0.05,桡神经在青、中、老三组CSA相比P>0.05。桡神经横截面积与身高、体重呈正相关性。桡神经横截面积与体重的最大相关系数是0.36(P<0.01),与身高的最大相关性系数是0.38(P<0.01)。结论桡神经在不同测量点其正常值不同,CSAs在性别对比中存在差异,青、中、老三组对比CSAs无差异,身高、体重与神经的大小呈正相关性。     

Ultrasonographic reference values for assessing normal radial nerve ultrasonography in the normal population

High-resolution ultrasound has been used recently to characterize median and ulnar nerves,but is seldom used to characterize radial nerves.The radial nerve is more frequently involved in entrapment syndromes than the ulnar and median nerves.However,the reference standard for normal radial nerves has not been established.Thus,this study measured the cross-sectional areas of radial nerves of 200 healthy male or female volunteers,aged 18 to 75,using high-resolution ultrasound.The results showed that mean cross-sectional areas of radial nerves at 4 cm upon the lateral epicondyle of the humerus and mid-humerus(midpoint between the elbow crease and axilla) were 5.14 ± 1.24 and 5.08 ± 1.23 mm2,respectively.The age and the dominant side did not affect the results,but the above-mentioned cross-sectional areas were larger in males(5.31 ± 1.25 and 5.19 ± 1.23 mm2) than in females(4.93 ± 1.21 and 4.93 ± 1.23 mm2,respectively).In addition,the cross-sectional areas of radial nerves were positively correlated with height and weight(r = 0.38,0.36,respectively,both P 0.05).These data provide basic clinical data for the use of high-resolution ultrasound for the future diagnosis,treatment,and prognostic evaluation of peripheral neuropathies.     

Development of a time-resolved immunofluorometric assay for quantifying platelet-derived microparticles in human plasma

Platelet-derived microparticles (PMPs) are considered a marker of platelet activation. They vary considerably in size, and flow cytometry, the predominant method used to assay PMPs, is only detecting larger PMPs (> 0.1 μm).

We describe here a method that quantifies the amount of PMP-located GPIIb antigen in detergent-treated platelet-free plasma (PPP) by means of a one-step time-resolved immunofluorometric assay (TR-IFMA). This assay uses a streptavidin-coated microwell plate and two different monoclonal antibodies to GPIIb (CD41), one conjugated to biotin and the other labeled with europium ion. A wide linear range standard curve with low background and a high sensitivity was obtained. Pre-assay ultracentrifugation or filtration of PPP extensively reduced the fluorometric signal, indicating that the GPIIb antigen is mainly particle-located. A strong correlation between the amount of GPIIb and PMP as detected by flow cytometry was found. Consequently, the assay can be used to study PMP-related phenomena and, in contrast to flow cytometry, can be used on frozen samples and is independent of PMP size.     

Hand-held myometry: reference values.

In thirteen major muscle groups of 50 healthy females and 50 males, aged 20-60 years, maximum voluntary contraction was measured with a hand-held dynamometer. The intrasession variation, the left-right variation, and the fifth and fiftieth centile values were calculated. The ratio of two observations within one session ranged from 0.85 to 1.18 and the ratio of left to right ranged from 0.82 to 1.22 (95% reference limits). In 20 volunteers the repeatability was tested after one week. The ratio of averages of three measurements in two successive weeks ranged from 0.82 to 1.23 (95% reference limits). There were only small differences between muscle groups concerning these ratios. A significant relation with age and weight/Quetelet Index could be demonstrated in some muscle groups. The mean strength of females is approximately two thirds of the strength of males. The data may be useful as reference values in the application of hand-held myometry.     

Sensitive and quantitative, 10-min immunofluorometric assay for D-Dimer in whole blood

INTRODUCTION: Normal concentrations of D-Dimer can be used to exclude venous thromboembolism (VTE). However, methods for sensitive and quantitative D-Dimer measurements at the point-of-care (POC) are still limited. MATERIALS AND METHODS: We developed a 10-min, non-competitive immunofluorometric assay for D-Dimer in citrated whole blood and plasma using pre-dispensed reagents dried in single assay wells. The simple, automated assay procedure comprises a 1:50 sample dilution, one-step incubation, washing, and time-resolved fluorometric measurement directly from the wet well surface. RESULTS: The limits of detection (background + 3SD) and quantification (CV <15%) were 0.05 and 0.2 mg/L D-Dimer, respectively, and the assay was linear up to 400 mg/L. Correlations to Roche TinaQuant (r=0.726, n=200) and Biopool Auto.Dimer (r=0.190, n=149) were carried out using citrated plasma. Diagnostic sensitivity, specificity, and negative (NPV) and positive (PPV) predictive values were 98.7%, 64.4%, 99.1% and 55.1%, and 92.2%, 81.0%, 95.9% and 68.3%, respectively, using cut-off values of 0.6 and 1.0 mg/L, respectively, in outpatients with deep vein thrombosis (DVT) and/or pulmonary embolism (PE) (n=77) compared with outpatients with various other diseases (n=174). The within- and between-run CVs near the cut-off values were < or =10% in both whole blood and plasma. The 95th percentile upper range in apparently healthy individuals was 0.68 mg/L of whole blood (n=101). CONCLUSIONS: The high sensitivity and NPV suggest that the rapid immunofluorometric assay could be valuable for rapid exclusion of VTE in outpatients. With appropriate cut-offs, the assay could potentially be used as a stand-alone test or combined with clinical probability assessment, but further studies are required.     

Normal controls and biological reference values in child psychiatry: defining normal

About half of adults volunteering as normal control research subjects may be rejected because of significant psychopathology, but no parallel study has been done to date for pediatric subjects. Of 152 applicants (ages 6 to 18) for participation as paid normal controls, 44% were found ineligible and at least 31.8% of the child volunteers had probable or certain psychiatric disorders. Successive screenings, including rating scales and structured interviews, were necessary to obtain controls meeting a defined standard of psychiatric health. Careful scrutiny of child volunteers in biological psychiatric research is needed to assure meaningful comparisons.     

Isoelectric focusing and crossed immunoelectrofocusing of CSF immunoglobulins in MS

Summary The 3 main Ig classes and the presence of free light chains were studied by isoelectric focusing and crossed immunoelectrofocusing in 100 CSF samples from patients with clinically definite or probable MS. Minute quantities of IgM and free light-chain (mostly lambda) components were found in 2 out of 11 and 5 out of 14 samples respectively. IgG and IgA were detected in all samples examined for these proteins and were found in the pI-ranges of 5.3–9.8 and 4.6–6.4 pH-units respectively. The gammaglobulin abnormalities found on isoelectric focusing were identified as microheterogeneous, oligoclonal IgG with predominantly kappa light-chain determinants. The IgG immunoprecipitates differed from those of normal subjects and the major abnormal components most frequently exhibited pI-values >8 pH-units. The IgA immunoprecipitates had 2–4 main components with some tendency to discontinuous subfractionation. This Ig class, however, did not exhibit the marked tendency to oligoclonal distribution found for IgG.

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Zusammenfassung Die 3 wichtigsten Ig-Klassen und das Vorhandensein von freien, leichten Ketten wurden durch Isoelektrofokusierung und durch gekreuzte Immuno-Elektrofokusierung im Liquor von 100 Patienten mit klinisch gesicherter oder wahrscheinlicher MS untersucht. Sehr kleine Mengen von IgM und von freien, leichten Kettenkomponenten (vor allem lambda) wurden in 2 von 11, respektive in 5 von 14 Fällen gefunden. IgG und IgA wurden in allen Fällen, in welchen nach diesen Proteinen gesucht wurde, im pI-Bereich von 5.3–9.8, respektive 4.6–6.4 pH-Einheiten gefunden. Die von Isoelektrofokusierung vorgefundenen Gammaglobulinanomalien wurden als mikroheterogene, oligoklonale IgG mit leichten Ketten vom überwiegend Kappa-Typ beschrieben. Die IgG-Immunoprezipitate unterschieden sich von normalen Fällen; die größeren abnormen Komponenten wiesen häufig pI-Werte von >8 pH-Einheiten auf. Die IgA-Immunoprezipitate hatten 2–4 Hauptkomponenten mit Tendenz zu unterbrochener Subfraktionierung. Diese Ig-Klasse wies jedoch nicht die bei IgG vorgefundene ausgesprochene Neigung zu oligoklonaler Verteilung auf.

     

Cross-sectional area reference values for nerve ultrasonography

Ultrasound allows for a non-invasive structural assessment of nerves, muscles, and surrounding tissues, and therefore it is increasingly being used as a supplement to traditional electrodiagnostic studies. As investigators have begun to use ultrasound to explore peripheral nerves, it has become clear that conditions such as entrapment, hereditary neuropathies, acquired neuropathies, trauma, and nerve tumors result in an increase in nerve cross-sectional area. Reference values have not been published for the cross-sectional area of many nerves commonly studied in diseases of the peripheral nervous system, so our goal was to obtain reference values for the nerve cross-sectional area at the following sites: radial at antecubital fossa; radial at distal spiral groove; musculocutaneous in upper arm; trunks of the brachial plexus; vagus at carotid bifurcation; sciatic in distal thigh; tibial in popliteal fossa; tibial in proximal calf; tibial at ankle; peroneal in popliteal fossa; peroneal at fibular head; and sural in distal calf. Mean cross-sectional area, as well as side-to-side differences, are reported for each site, and qualitative data are provided to guide imaging at each site. The information provided in this study should serve as the starting point for quantitatively evaluating these nerve sites with ultrasound.     

Sensitized immunofixation: a new technique for analyzing the oligoclonal pattern of CSF immunoglobulins

Intrathecal immunoglobulin synthesis is observed in more than 90% of all cases of multiple sclerosis, producing a specific CSF IgG oligoclonal electrophoretic pattern. The consensual method used as reference is isoelectric focusing (IEF). We developed a new CSF Ig analysis method by immunofixation (IF). The method includes an immunoenzymatic detection step performed directly on the gel allowing the use of unconcentrated CSF and avoiding the blotting step. The reliability of this method was established by the analysis of 210 CSF/serum pairs including defined, probable and possible MS, other inflammatory CNS diseases and controls (noninflammatory CNS diseases and peripheral nervous system diseases). Intrathecal IgG synthesis was detected in 95.5% of defined MS cases. The specificity for CNS inflammatory diseases including MS diagnosis, evaluated by comparison with controls, was 98.8%. This new method is quicker and visual interpretation is easier than with IEF. It is a semi-automated method that should be considered for standardization of CSF IgG analysis.     

Multiple sclerosis: rat and human oligodendrocytes are not the target for CSF immunoglobulins

CSF Ig from 28 patients with MS and 25 patients with other neurologic diseases (OND) were examined for their capacity to bind to rat or human oligodendrocytes in culture. We have used a double-label approach combining CSF Ig with antibodies against galactocerebroside (GalC). As normal human IgG at a concentration higher than 250 micrograms/ml were found to bind nonspecifically to oligodendrocytes in culture, patients' CSFs were used unconcentrated, at a final IgG concentration never exceeding 115 micrograms/ml. In these experimental conditions, we have not been able to detect any fixation of CSF Ig from MS (or OND) patients to rat or human GalC+ oligodendrocytes.     

DNA-labelled cytidine assay for the quantification of CAG repeats

The sequencing procedure has been used to determine the size of the CAG repeat expansion for the diagnosis of genetic disorders. Likewise, standard polymerase chain reaction (PCR) and gel electrophoresis techniques are applied for screening large number of patients. The trinucleotide repeats (TNR) region amplification by means of the PCR procedure was initially performed using 32-P end-labelled primers and currently carried out with fluorescently end-labelled primers. The goal to obtain reliable TNR quantification assays, at low cost and short assay times, represents a challenge for the molecular diagnosis aimed at massive screening of affected populations. In the current work, we obtained preliminary results of a new methodology for the detection and size estimation of CAG expanded alleles. The assay was based on an indirect enzyme linked immunosorbent assay (ELISA) for quantifying the amount of labelled cytidines in DNA molecules. The label, 6-(p-bromobenzamido)caproyl radical, was introduced by the transamination and acylation reactions. A group of model sequences containing different numbers of CAG repeats, as well as the ATXN3 (ataxin 3) gene (from subjects suffering type 3 spinocerebellar ataxia SCA3) were used for assay standardization. The assay is simple, inexpensive, and easy to perform and differentiates distinct degrees of CAG expansions.     

Dorsal and lumbar paraspinal electromyographic study. Multi-MUP analysis and drawing up normal values in a reference population]

OBJECTIVE: The aim of this study was to contribute to draw up reference values relating to electromyographic (EMG) parameters in dorsal and lumbar paraspinal muscles. MATERIALS AND METHODS: 75 healthy subjects without back pain underwent electromyography of multifidus bundles, which are innervated uni-segmentally by the dorsal ram of the spinal nerve. T8, L3, L4, L5 and S1 myotomes were systematically explored. Output variables were spontaneous denervation activity and quantitative EMG data obtained by multi-MUP (Motor Unit action Potential) analysis. RESULTS: No abnormal insertional or spontaneous activity (such as fibrillation, positive sharp waves or fasciculation) was recorded at rest. Neither sex nor age influenced motor unit action potential features in our series. Reference values were drawn up for T8 and L5 segmental levels using the mean values of 20 motor unit potentials in each patient studied, the reference interval being defined by the lower and the upper outlier limits on individual values. CONCLUSION: This study offers reference data to electromyographers to help better identifying possible myogenic or neurogenic pathological changes, especially in lumbosacral radiculopathies.     

Suspected and clinically definite multiple sclerosis: the relationship between CSF immunoglobulins and clinical course.

CSF immunoglobulins were examined in 103 patients with clinically definite multiple sclerosis, 106 patients with either suspected or progressive possible multiple sclerosis and 72 patients with other neurological diseases. Raised CSF IgG index and oligoclonal banding were found in 71% and 75% of clinically definite multiple sclerosis patients respectively and both tests were abnormal in 11% of patients with other neurological diseases. The CSF IgG index and the presence of oligoclonal IgG did not relate to the severity or duration of established disease in these patients. In patients with suspected and progressive possible multiple sclerosis, both a raised IgG index and the presence of oligoclonal banding were found significantly more frequently than in the OND group. Abnormalities of these parameters were significantly correlated with the presence of an abnormal evoked response in these patients (chi 2 = 10.16 p less than 0.01). When 47 patients with suspected multiple sclerosis were studied prospectively the presence of oligoclonal banding at presentation was associated with development of further disease activity.     

CA-1 method, a novel assay for quantification of normal prothrombin using a Ca2+ -dependent prothrombin activator, carinactivase-1.

We established a novel prothrombin assay, designated CA-1 method, for quantification of normal prothrombin in application of a Ca2+ -dependent prothrombin activator, carinactivase-1 (CA-1), found in the venom of Echis carinatus leucogaster. On microplate, thrombin converted from normal prothrombin in plasma sample by CA-1 cleaves a thrombin specific chromogenic substrate, t-butoxy-Val-Pro-Arg-p-nitroanilide and liberates p-nitroaniline. Then, the normal prothrombin level is decided by measuring the velocity of p-nitroaniline liberation. Normal prothrombin levels in plasma from warfarin-treated individuals were highly correlated with coagulant activities assayed by both prothrombin time and thrombotest. CA-1 method is not only a rapid and highly sensitive chromogenic microplate assay for quantification of normal prothrombin in the range of 10-200 ng/100 microl in plasma samples but also suitable for analyses of many samples in a short time. In addition, normal prothrombin levels obtained by CA-1 method are not inhibited by EDTA and heparin, which reduce prothrombin time and thrombotest activities. CA-1 method is a novel assay for monitoring coagulant activity in warfarin-treated individuals.