AHI1 gene mutation in a consanguineous Iranian family affected by Joubert syndrome: A case report

This point of detected mutation could be considered as a novel mutational hotspot point that carried in patient ancestors. Moreover, the obtained results and family history suggest a precise genetic consulting and molecular prenatal evaluation for suspect individuals with a family history of mental and physical abnormalities.     

COVID‐19 spreading across world correlates with C677T allele of the methylenetetrahydrofolate reductase (MTHFR) gene prevalence

BackgroundHomocysteine assessment has been proposed as a potential predictive biomarker for the severity of COVID‐19 infection. The purpose of this review was to analyze the correlation between the prevalence of MTHFR C677 T gene polymorphism and COVID‐19 incidence and mortality worldwide.MethodsData regarding MTHFR C677 T gene mutation were obtained from the interrogation of the Genome Aggregation Database (genomAD), which is publicly available from the web“https://gnomad.broadinstitute.org.” COVID‐19 cases, including prevalence and mortality, were obtained from“https://www.worldometers.info/coronavirus” 27 August 2020.ResultsThere is a clear trend toward the worldwide prevalence of MTHFR 677 T and COVID‐19 incidence and mortality. The prevalence of MTHFR 677 T allele in the Latino population, and the incidence and mortality for COVID‐19 was higher for this ethnic group than that reported for most other populations globally. Statistical analysis showed a relatively strong correlation between C677 T and death from coronavirus.ConclusionsGenetic polymorphism of MTHFR C677 T may modulate the incidence and severity of COVID‐19 pandemic infection.     

Subcutaneous panniculitis‐like T‐cell lymphoma in a young girl presenting with periorbital edema and fever: A case report

Subcutaneous panniculitis‐like T‐cell lymphoma is a rare and highly malignant extra‐nodal lymphoma. It has a wide range of clinical presentations (such as periorbital swelling as in our case) and should be considered in the differential diagnosis of systemic lupus erythematosus, especially in children.     

Identification of a compound heterozygous missense mutation in LAMA2 gene from a patient with merosin‐deficient congenital muscular dystrophy type 1A

BackgroundMerosin‐deficient congenital muscular dystrophy type 1A (MDC1A) is occurred by mutations in LAMA2 gene that encodes the laminin α2 chain (merosin). MDC1A is a predominant subtype of congenital muscular dystrophy. Herein, we identified two missense mutations in LAMA2 gene in compound heterozygous status in an Iranian patient with MDC1A using whole‐exome sequencing (WES).MethodsIn the present study, we evaluated genetic alterations in an Iranian 35‐month‐old boy with MDC1A and his healthy family using WES method. The identified mutations further confirmed by Sanger sequencing method. Finally, in silico analysis was conducted to further evaluation of molecular function of the identified genetic variants.ResultsWe identified two potentially pathogenic missense mutations in compound heterozygous state (c.7681G>A p.Gly2561Ser and c.4840A>G p.Asn1614Asp) in LAMA2 gene as contributing to the MDC1A phenotype. The healthy parents of our proband are single heterozygous for identified mutations. These variants were found to be pathogenic by in silico analysis.ConclusionsIn general, we successfully identified LAMA2 gene mutations in an Iranian patient with MDC1A using WES. The identified mutations in LAMA2 gene can be useful in genetic counseling, prenatal diagnosis, and predicting prognosis of MDC1A.     

Sequential allogeneic transplantation and ruxolitinib maintenance for a synchronous PCM1‐JAK2 positive myeloid sarcoma and acute B‐lymphoblastic leukemia

The translocation t(8;9)(p22;p24) results in the production of a chimeric PCM1‐JAK2 fusion protein leading to the constitutive activation of the Janus Kinase 2 that renders this disease potentially sensitive to ruxolitinib. Here, we report an interesting case of PCM1‐JAK2 myeloproliferative neoplasm evolving in myeloid sarcoma and B precursor ALL.     

Chorioamnionitis and neonatal sepsis due to extended‐spectrum beta‐lactamase‐producing Escherichia coli infection: a case report

Chorioamnionitis is an acute inflammation of the membranes and chorion of the placenta typically due to ascending polymicrobial infection in the setting of membrane rupture. It is a common complication of pregnancy associated with significant maternal, perinatal, and long‐term adverse outcomes. We present a case of placental infection leading to preterm delivery, severe neonatal sepsis, maternal wound infection, postnatal readmission, and prolonged hospital stay. This virulent infection was caused by multidrug–resistant extended‐spectrum beta‐lactamase (ESBL)‐producing Escherichia Coli (E. Coli), which represent a major worldwide threat according to the Centre for Disease Control and Prevention (CDC). It was managed with appropriate antibiotic therapy, patient‐centered approach, and multidisciplinary team involvement that led to favourable maternal and neonatal outcome.     

Perampanel markedly improved clinical seizures in a patient with a Rett‐like phenotype and 960‐kb deletion on chromosome 9q34.11 including the STXBP1

Intractable epilepsy was successfully controlled using perampanel, an α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole propionic acid‐type glutamate receptor antagonist, in a 27‐year‐old woman who presented with a Rett syndrome‐like phenotype and novel 960‐kb deletion involving syntaxin‐binding protein 1 on chromosome 9q34.11. Perampanel may be an effective antiepileptic drug for intractable epilepsy associated with STXBP1 mutations.     

Simultaneous disruption of circulating miR‐21 and cytotoxic T lymphocytes (CTLs): Prospective diagnostic and prognostic markers for esophageal squamous cell carcinoma (ESCC)

Background Esophageal squamous cell carcinoma (ESCC) as the most prominent type of esophageal cancer (EC) in developing countries encompasses a substantial contribution of cancer‐related mortalities and morbidities. Cytotoxic T lymphocytes (CTLs) are the major subset of effector T cells against cancer. However, the microRNAs involved in the development and regulation of CTLs could be disrupted in cancers such as EC.MethodsHere, we evaluated the population of IL‐10, TGF‐β, IFN‐γ, and IL‐17a‐producing CD3+CD8+ T cells, their association with the circulating levels of miR‐21 and miR‐29b, and their diagnostic and/or prognostic (after 160 weeks of follow‐up) utilities in 34 ESCC patients (12 newly diagnosed: ND, 24 under‐treatment: UT) and 34 matched healthy donors.ResultsThe population of IL‐10 and TGF‐β‐producing CTLs (CD8+ Tregs) were considerably expanded, in addition to the overexpression of miR‐21 in both groups (ND and UT) of ESCC patients, while the frequency of Tc17 and CD8+ Treg cells increased only in UT patients. The expression means of TGF‐β and IL‐10 in CTLs were considered to be excellent biomarkers (1 ≥ area under the curve: AUC ≥0.9) in distinguishing ESCC patients and associated subgroups from healthy subjects. Moreover, the lower expressions of TGF‐β, IL‐17a, IL‐10, and IFN‐γ in CTLs were associated with ESCC better prognosis.ConclusionsThe association between the impaired function of CD3+ CD8+ T cell subsets and miR‐21 expression could be introduced as novel therapeutic targets and powerful diagnostic and prognostic markers for ESCC.     

Genotyping,antifungal susceptibility,enzymatic activity,and phenotypic variation in Candida albicans from esophageal candidiasis

BackgroundEsophageal candidiasis is the most frequent form of esophagitis. The pathogenicity of Candida spp. is related to a combination of microbial factors, hydrolytic enzyme secretion and phenotypic switching. This study was designed to investigate esophageal candidiasis, antifungal activity, enzymatic activity patterns, phenotyping, and genotyping profiles of Candida albicans species.MethodsNine hundred thirty‐three visited patients were evaluated, and esophageal biopsies from patients were included in this study during 2019–2020. Direct smear, Gram staining, and culture on CHROMagar were performed for each sample. Isolated species were identified with conventional procedures and PCR‐RFLP. Susceptibility to antifungals was determined according to CLSI guidelines. ABC typing, phenotype switching, hemolysin, proteinase, phospholipase, and esterase activity were also determined with the appropriate protocols.ResultsTwenty‐three (2.5%) patients (mean age 55.2 years) were diagnosed with esophageal candidiasis. The species isolated were 19(82.6%) C. albicans, 3(13.1%) C. glabrata, and 1(4.3%) C. tropicalis. Genotype A (57.9%) was the predominant type in C. albicans isolates. 50% of C. albicans isolates exhibited a white phenotype. A high level of phospholipase (47.4%), hemolysin (68.4%), and proteinase activity (36.8%) was observed in the C. albicans isolates. Only three C. glabrata isolates displayed non‐wild type susceptibility to voriconazole and itraconazole.ConclusionThis study shows that C. albicans are still the most frequent isolates from patients with esophageal candidiasis. The predominance of genotype A, the white phenotype, and strong hemolysin activity may indicate a high prevalence of pathogenicity in these isolates. Sensitivity to antifungal drugs was greatest for amphotericin and fluconazole.     

Bilateral posterior lamellar corneal transplant surgery in an infant of 17 weeks old: Surgical challenges and the added value of intraoperative optical coherence tomography

This study aimed to describe the surgical challenges, management, and value of intraoperative optical coherence tomography in a case of a bilateral Descemet Stripping Automated Endothelial Keratoplasty corneal transplantation at 17 weeks of age for the treatment of severe posterior polymorphous corneal dystrophy resulting from a de novo mutation of the OVOL2‐gene.     

A novel Tc‐99 m and fluorescence labeled peptide as a multimodal imaging agent for targeting angiogenesis in a murine tumor model

The serine–aspartic acid–valine (SDV) peptide binds specifically to integrin α_Vβ₃. In the present study, we successfully developed a TAMRA–GHEG–ECG–SDV peptide labeled with both Tc‐99 m and TAMRA to target the integrin α_Vβ₃ of tumor cells; furthermore, we evaluated the diagnostic performance of Tc‐99 m TAMRA–GHEG–ECG–SDV as a dual‐modality imaging agent for tumor of the murine model. TAMRA–GHEG–ECG–SDV was synthesized using Fmoc solid‐phase peptide synthesis. Radiolabeling of TAMRA–GHEG–ECG–SDV with Tc‐99 m was done using ligand exchange methods. Labeling stability and cytotoxicity studies were performed. Gamma camera imaging, biodistribution and ex vivo imaging studies were performed in murine models with HT‐1080 and HT‐29 tumors. A tumor tissue slide was prepared and analyzed using confocal microscopy. After radiolabeling procedures with Tc‐99 m, the Tc‐99 m TAMRA–GHEG–ECG–SDV complexes were prepared in high yield (>99%). In the gamma camera imaging study, a substantial uptake of Tc‐99 m TAMRA–GHEG–ECG–SDV into HT‐1080 tumor (integrin α_Vβ₃ positive) and low uptake of Tc‐99 m TAMRA–GHEG–ECG–SDV into HT‐29 tumor (integrin α_Vβ₃ negative) were demonstrated. A competition study revealed that HT‐1080 tumor uptake was effectively blocked by the co‐injection of an excess concentration of SDV. Specific uptake of Tc‐99 m TAMRA–GHEG–ECG–SDV was confirmed by biodistribution, ex vivo imaging and confocal microscopy studies. Our in vivo and in vitro studies revealed substantial uptake of Tc‐99 m TAMRA–GHEG–ECG–SDV in the integrin α_Vβ₃‐positive tumor. Tc‐99 m TAMRA–GHEG–ECG–SDV could be a good candidate for a dual‐modality imaging agent targeting tumor angiogenesis. Copyright © 2016 John Wiley & Sons, Ltd.     

Downregulation of Talin‐1 is associated with the increased expression of miR‐182‐5p and miR‐9‐5p in coronary artery disease

BackgroundEvidence indicates that the dysregulation of extracellular matrix (ECM) components can lead to cardiovascular diseases. The Talin‐1 (TLN1) gene is a major component of the ECM, and it mediates integrin adhesion to the ECM. In this study, we aimed to determine microRNAs (miRs) that regulate the expression of TLN1 and determine expression alterations in TLN1 and its targeting miRs in coronary artery disease (CAD).MethodsData sets of CAD and normal samples of blood exosomes were downloaded, and TLN1 was chosen as one of the genes with differential expressions in an in silico analysis. Next, miR‐182‐5p and miR‐9‐5p, which have a binding site on 3´‐UTR of TLN1, were selected using bioinformatics tools. Then, the miR target site was cloned in the psiCHECK‐2 vector, and direct interaction between the miR target site and the TLN1 3′‐UTR putative target site was investigated by luciferase assay. The expression of miR‐182‐5p, miR‐9‐5p, and TLN1 in the serum samples of CAD and non‐CAD individuals was assessed via a real‐time quantitative polymerase chain reaction.ResultsOur data revealed that miR‐182‐5p directly regulated the expression of TLN1. Moreover, miR‐182‐5p and miR‐9‐5p were significantly upregulated in the CAD group. Hence, both bioinformatics and experimental analyses determined the downregulated expression of TLN1 in the CAD samples.ConclusionsOur findings demonstrated that miR‐182‐5p and miR‐9‐5p could play significant roles in TLN1 regulation and participate in CAD development by targeting TLN1. These findings introduce novel biomarkers with a potential role in CAD pathogenesis.     

Genetic research and clinical analysis of β‐globin gene cluster deletions in the Chinese population of Fujian province: A 14‐year single‐center experience

BackgroundHeterozygotes of HPFH and δβ thalassemia are clinically asymptomatic or have mild hemoglobin (Hb) values. However, when both HPFH and δβ‐thalassemia are coinherited with heterozygous β‐thalassemia, patients may progress to a clinical phenotype of thalassemia intermedia or thalassemia major. The purpose of this study was to characterize the genotypes and analyze the phenotypes of these disorders in Fujian Province, to offer advice for genetic counseling and accurate prenatal diagnosis in this region. A total of 55 001 subjects were participated in thalassemia screening. 142 subjects with HbF levels ≥10%, before the blood transfusion, were selected for further investigation.MethodsMultiplex ligation‐dependent probe amplification (MLPA) and Gap‐PCR were used to screen for three β‐globin gene cluster deletions: Chinese ^Gγ(^Aγδβ)⁰ thalassemia and Southeast Asia HPFH (SEA‐HPFH) deletion and 1357 bp deletion (NG‐000007.3:g.69997‐71353 del 1357).ResultsA total of 142 patients with HbF (≥10%) were enrolled to characterize the molecular basis of β‐globin gene cluster deletions in our study; 22 cases 0.04% (22/55 001) were definitively diagnosed with β‐globin gene cluster deletions. Ten cases were heterozygous for the Chinese ^Gγ(^Aγδβ)⁰‐thal mutations, 10 cases were heterozygous for SEA‐HPFH, and one case was compound heterozygous for SEA‐HPFH and the α‐thal mutation. The 1357 bp deletion (NG‐000007.3:g.69997‐71353 del 1357) was detected in one case. Moreover, the hemoglobin A₂ levels in patients who were heterozygous for Chinese ^Gγ(^Aγδβ)⁰‐thal were statistically lower than in cases with SEA‐HPFH deletion(p < 0.05).ConclusionIn Fujian Province, the prevalence of common β‐globin gene cluster deletions was 0.04%. What''s more, the most common β‐globin cluster deletions are the Chinese ^Gγ(^Aγδβ)⁰ and SEA‐HPFH.     

Correlation analysis between vitamin A,D, and E status with altitude,seasonal variation,and other factors,among children aged 0–6 years in a Chinese population living in the Tibetan plateau of Ganzi prefecture

ObjectiveWe attempted to understand the status of vitamin (Vit) A, D, and E in children aged 0–6 living in the Tibetan plateau areas of Ganzi prefecture, to provide the basis for relevant government departments to carry out physical examinations of these children and to prevent and cure four key diseases (Infantile diarrhea, nutritional anemia, rickets, and infantile pneumonia).MethodsSerum retinol and tocopherol levels were detected using high‐performance liquid chromatography (HPLC). Serum levels of 25‐(OH)D were detected by high‐performance liquid chromatography–tandem mass spectrometry (LC–MS). The polynomial logistic regression was used to analyze the effects of age, season, altitude, and gender on Vit A, D, and E levels.ResultsVit A and D had the lowest mean serum levels before the age of 1 year and with the most significant deficiency rates. The lowest Vit E levels were seen in the Toddlerhood group. The rates of deficiency and insufficiency were the highest. Vit A, D, and E levels were significantly affected by seasonal changes and were significantly higher in the summer than in any other season. Vit A and D were significantly affected by altitude, and their levels were lowest above 4 km.ConclusionThe overall levels of Vit A, D, and E in children aged 0–6 in the Tibetan plateau areas of Ganzi prefecture were lower than those in the plain''s areas.     

Efficacy and safety of three new oral antiviral treatment (molnupiravir,fluvoxamine and Paxlovid) for COVID-19：a meta-analysis

BackgroundThe coronavirus disease (COVID-19) epidemic has not been completely controlled. Although great achievements have been made in COVID-19 research and many antiviral drugs have shown good therapeutic effects against COVID-19, a simple oral antiviral drug for COVID-19 has not yet been developed. We conducted a meta-analysis to investigate the improvement in mortality or hospitalization rates and adverse events among COVID-19 patients with three new oral antivirals (including molnupiravir, fluvoxamine and Paxlovid).MethodsWe searched scientific and medical databases, such as PubMed, Web of Science, Embase and Cochrane Library for relevant articles and screened the references of retrieved studies on COVID-19.ResultsA total of eight studies were included in this study. The drug group included 2440 COVID-19 patients, including 54 patients who died or were hospitalized. The control group included a total of 2348 COVID-19 patients, including 118 patients who died or were hospitalized. The overall odds ratio (OR) of mortality or hospitalization was 0.33 (95% confidence interval [CI], 0.22–0.49) for COVID-19 patients in the drug group and placebo group, indicating that oral antiviral drugs were effective for COVID-19 patients and reduced the mortality or hospitalization by approximately 67%.ConclusionsThis study showed that three novel oral antivirals (molnupiravir, fluvoxamine and Paxlovid) are effective in reducing the mortality and hospitalization rates in patients with COVID-19. In addition, the three oral drugs did not increase the occurrence of adverse events, thus exhibiting good overall safety. These three oral antiviral drugs are still being studied, and the available data suggest that they will bring new hope for COVID-19 recovery and have the potential to be a breakthrough and very promising treatment for COVID-19.

KEY MESSAGES

• Many antiviral drugs have shown good therapeutic effects, and there is no simple oral antiviral drug for COVID-19 patients.
• Meta-analysis was conducted for three new oral antivirals to evaluate the improvement in mortality or hospitalization rates and adverse events among COVID-19 patients.
• We focussed on three new oral Coronavirus agents (molnupiravir, fluvoxamine and Paxlovid) and hope to provide guidance for the roll-out of oral antivirals.

     

内镜逆行胰胆管造影术与经皮肝穿刺胆道引流术治疗肝外胆管梗阻性黄疸的对比研究

目的探讨内镜逆行胰胆管造影术(ERCP)与经皮肝穿刺胆道引流术(PTCD)在治疗良、恶性肝外胆管梗阻性黄疸方面各自的优劣。方法回顾性分析2013年5月至2014年7月期间兰州大学第二医院普外四科收治的肝外胆管梗阻性黄疸患者并分别采用ERCP与PTCD治疗的临床资料,比较2种方法的手术成功率、5 d血清胆红素下降水平及术后常见并发症的发生率等。结果与PTCD相比,ERCP在治疗良性肝外胆管梗阻性黄疸时,术后5 d血清总胆红素水平下降速度较快[(94.9±11.58)μmol/L vs.(84.3±15.50)μmol/L,t=2.946,P=0.005],术后并发症发生率较低(3.3%vs.21.4%,χ2=4.469,P=0.035),但手术成功率(96.7%vs.92.9%,χ2=0.429,P=0.513)二者无统计学差异。在治疗恶性肝外胆管梗阻性黄疸时,两种方法术后常见并发症的发生率(9.5%vs.18.5%,χ2=0.767,P=0.381)虽无明显差异,但相比ERCP,在手术成功率(95.2%vs.70.4%,χ2=4.795,P=0.029)和术后5 d血清总胆红素水平下降速度[(206.3±13.26)μmol/L vs.(186.8±20.59)μmol/L,t=-2.516,P=0.015]方面PTCD组效果较为显著。结论对于良性肝外胆管梗阻性黄疸患者ERCP不仅具有创伤小、并发症少、起效快、疗效好的优点,而且可以从根本上解除梗阻;而对于恶性肝外胆管梗阻性黄疸,由于PTCD手术成功率高,可以有效缓解梗阻症状,快速降低血清胆红素水平,改善肝功能,提高患者生活质量,因此成为首选方法。     

Serum concentrations of small dense low‐density lipoprotein cholesterol and lipoprotein(a) are related to coronary arteriostenosis in Takayasu arteritis

BackgroundSerum small dense low‐density lipoprotein cholesterol (sdLDL‐C) and lipoprotein(a) [Lp(a)] levels are related to coronary disease, but their specific associations with coronary arteriostenosis in Takayasu arteritis (TA) have not been ascertained. This study explored the correlations between serum sdLDL‐C and Lp(a) levels and coronary arteriostenosis in TA patients as well as the degree of artery stenosis.MethodsThis retrospective study included 190 TA patients and 154 healthy subjects. TA patients were divided into three categories based on the degree of coronary stenosis: Group I, stenosis >50%; Group II, stenosis 1%–50%; and Group III, stenosis 0%. Independent risk factors for coronary arteriostenosis in TA were identified by logistic regression, followed by receiver operating characteristic curve analysis to determine the specificity and sensitivity of risk factors and Youden''s Index score calculation to determine the cutoff points.ResultsTakayasu arteritis patients had significantly higher serum levels of sdLDL‐C and Lp(a) than healthy controls (p < 0.0001). The total cholesterol, triglyceride, LDL‐C, sdLDL‐C, and Lp(a) levels and the sdLDL‐C/LDL‐C ratio in Group I were significantly higher than those in Groups II and III (p < 0.05). An elevated serum sdLDL‐C level elevated the risk of coronary arteriostenosis by 5‐fold (cutoff value, 0.605 mmol/l). An increased serum Lp(a) level increased the risk of coronary arteriostenosis by 3.9‐fold (cutoff value, 0.045 g/l). An elevated sdLDL‐C/LDL‐C ratio increased the risk of coronary arteriostenosis by 2.1‐fold (cutoff value, 0.258).ConclusionsSerum sdLDL‐C and Lp(a) levels and sdLDL‐C/LDL‐C ratio may be used as diagnostic factors for coronary arteriostenosis in TA patients.     

A New Monoclonal Antibody,mAb 4A12, Identifies a Role for the Glycosaminoglycan (GAG) Binding Domain of RANTES in the Antiviral Effect against HIV-1 and Intracellular Ca2+ Signaling

The β-chemokine RANTES (regulated on activation, normal T cell expressed and secreted) suppresses the infection of susceptible host cells by macrophage tropic strains of HIV-1. This effect is attributed to interactions of this chemokine with a 7-transmembrane domain receptor, CCR5, that is required for virus–cell fusion and entry. Here we identify domains of RANTES that contribute to its biological activities through structure–function studies using a new monoclonal antibody, mAb 4A12, isolated from mice immunized with recombinant human RANTES. This monoclonal antibody (mAb) blocked the antiviral activity of RANTES in infectivity assays with HIV-1_Bal, and inhibited the mobilization of intracellular Ca²⁺ elicited by RANTES, yet recognized this chemokine bound to cell surfaces. Epitope mapping using limited proteolysis, reversed phase high-performance liquid chromatography, and mass spectrometry suggest that residues 55–66 of RANTES, which include the COOH-terminal α-helical region implicated as the glycosaminoglycan (GAG) binding domain, overlap the determinant recognized by mAb 4A12. This is supported by affinity chromatography studies, which showed that RANTES could be eluted specifically by heparin from a mAb 4A12 immunoaffinity matrix. Removal of cell surface GAGs by enzymatic digestion greatly reduced the ability of mAb 4A12 to detect RANTES passively bound on cell surfaces and abrogated the ability of RANTES to elicit an intracellular Ca²⁺ signal. Taken together, these studies demonstrate that the COOH-terminal α-helical region of RANTES plays a key role in GAG-binding, antiviral activity, and intracellular Ca²⁺ signaling and support a model in which GAGs play a key role in the biological activities of this chemokine.     

Reference data for quantitative sensory testing (QST): refined stratification for age and a novel method for statistical comparison of group data

Clinical use of quantitative sensory testing (QST) requires standardization. The German research network on neuropathic pain (DFNS) solves this problem by defining reference data stratified for test site, gender and age for a standardized QST protocol. In this report we have targeted two further problems: how to adjust for age-related sensory changes, and how to compare groups of patients with the reference database. We applied a moving average across ages to define reference values per decade. This analysis revealed that women were more sensitive to heat pain independent of age. In contrast, functions were converging at older age for blunt pressure pain, but diverging for punctate mechanical pain (pin prick). The probability that an individual patient dataset is within the range of normal variability is calculated by z-transform using site-, gender- and age-specific reference data. To compare groups of patients with reference data, we evaluated two techniques: A: paired t-test versus fixed mean; i.e. the reference mean value is considered as the known population mean, B: non-paired t-test versus the reference dataset and number of cases restrained to the same number of cases as the patient data set. Simulations for various sample sizes and variances showed that method B was more conservative than method A. We present a simple way of calculating method B for data that have been z-normalized. This technique makes the DFNS reference data bank applicable for researchers beyond the DFNS community without a need for subsampling of subjects from the database.