bluuebunny/crossref_metadata_2025 huggingface dataset:
Qualitative Immunoglobulin Abnormalities in HIV-Positive Patients: Long-Term Follow-Up
Authors: Pontet Françoise , Gué Xavier , Mazeron Marie-Christine , Alain Sophie , Sanson-Le Pors Marie-José
Published: October 2004

We studied the immunoglobulins of a cohort of 212 HIV-positive patients followed-up for 13 years. The qualitative study of immunoglobulins by immunoelectrophoresis and immunofixation distinguished three groups of patients: those with monoclonal immunoglobulins, those with minor abnormalities of immunoglobulins and those with polyclonal immunoglobulins. We characterized these groups according to age, sex, immunoglobulin isotypes, and survival curves. The results show that this population of immunoglobulinopathies has distinctive characteristics. In particular, the presence of immunoglobulin abnormalities has no significant prognostic value.
The Human Genome Project: From Mapping to Sequencing
Authors: Weissenbach Jean
Published: October 2004

Until recently, the “human genome” programs were mainly directed towards the development of maps to identify disease genes. The genetic map comprises about 8000 highly informative second generation markers of the microsatellite type. The density of markers is now sufficient to localize a gene for a monogenic disease with a precision of 1 to 2 million base pairs easily, and to define intervals which contain susceptibility genes for multifactorial disorders. A third generation map based on single nucleotide polymorphisms that can be genotyped using DNA chip technology is in progress. The physical map, based on sets of overlapping yeast artificial chromosomes ordered using sequence-tagged sites, covers over 90 % of the genome. However, this physical map cannot serve as a support for sequencing because of the numerous rearrangements that occur in yeast artificial chromosomes. An international network of laboratories has mapped a set of more than 30000 expressed sequences from cDNAs using whole genome radiation hybrids that enable integration of genes within existing maps. The human genome program is now progressively shifting to massive sequencing, although sequence ready maps are not available for the major part of the human genome. Similarly, our capacity to interpret the available genomic sequence remains limited.
The Human Genome Project and the Role of Genetics in Health Care
Authors: van Ommen Gert-Jan B.
Published: October 2004

The Human Genome Project, the mapping of our 100000 genes and the sequencing of all of our DNA, will have major impact on biomedical resarch and the therapeutic and preventive health care. The tracing of genetic diseases to their molecular causes is rapidly expanding diagostic and preventive options, while the increased insights into molecular pathways open tremendous perspectives for pharmacological and genetic therapies. The design of animal model systems for the functional study of disease and development of bioinformatics and biostatistics to improve our pattern recognition abilities are greatly accelerating progress. However, the optimal value from the current explosion of ‘data mining’ possibilities will only be gained when the basic data are made and kept publicly accessible, at the same time preventing the jeopardisation of the protection of intellectual property, arising from downstream inventions. This is one of the goals of HUGO, the international Human Genome Organisation, established 9 years ago to assist coordinating data acquisition and exchange and societal implementation of the genome project. Additional points of major importance in this historic endeavour are the safe-guarding of a worldwide balance in the contribution and benefits to countries and population, the prevention of stigmatisation and discrimination of individuals and groups and the maintenance of respect for the priceless diversity of our world's cultures and traditons.
Population Genomics: Laying the Groundwork for Genetic Disease Modeling and Targeting
Authors: Gulcher Jeff , Stefansson Kari
Published: October 2004

The family has proven the most appropriate unit with which to study Mendelian diseases. There are, however, certain limitations on the use of the family as a fundamental unit in the study of common diseases, most of which are complex genetic diseases. The groups that are most likely to yield the genetics of complex diseases are isolated populations with strong founder effects. Therefore, access to such populations is proving to be a precious resource in the work on the genetics of common diseases. The Icelandic population is an excellent population for the study of the genetics of common diseases; it is genetically homogeneous, with founder effects for many traits, and the genealogy of the entire nation is well documented back to the founding days. Furthermore, the nature of the Icelandic national health care system facilitates the assignment of phenotypes in the search for disease genes.

Decode Genetics has begun to study of the genetics of 20 of the most common diseases in the Western parts of the world. The company has placed the groundwork for the construction of an encrypted database with information on the health care of the entire nation, genealogy of the entire nation, genotyping information with high density of markers on a large part of the nation (including typing for known disease genes), and resource use in the Icelandic health care system. The plan is to build the database with approval of participating individuals as well as Icelandic government and health care officials. The database will be used to model health care as viewed in the context of genetic predisposition to the development of disease. The database will also be used in the search for drug targets in complex diseases and in the solution of pharmacogenomic problems. Basing the company in Iceland directly benefits the population in terms of employment and return on investment as well as providing the health care system with an information resource which may be used in preventive medicine and in the optimization of health care in Iceland.
Signal Transduction Pathways and Modulation of Gene Activity
Authors: Weitzman Jonathan B. , Yaniv Moshe
Published: October 2004

Research over the last decade has provided us with much information about the mechanisms that integrate signal transduction pathways with specific gene expression programs. We discuss the types of mechanisms used by different pathways to modulate transcription factor activity, citing examples from diverse molecular systems. Careful regulation of these pathways is essential to maintain balanced transcriptional control. Understanding the mechanisms that control gene activity will enable us to intervene therapeutically in diseases associated with deregulated transcriptional activity.
Do Overexpressed Oncoproteins Cause Malignant Growth of Cancer Cells? – Studied by Antisense Oligonucleotides
Authors: Lin Shwu-Bin
Published: October 2004

Antisense oligonucleotides were used to demonstrate the importance of insulin-like growth factor II and α-fetoprotein for the growth of hepatoma cell lines. The level of insulin-like growth factor II was found to correlate positively with cell proliferative activity, whereas α-fetoprotein was not. We have developed an in vitro system for the screening of antisense oligonucleotides effective for inhibiting target protein production. Using this system, the effectiveness of antisense oligonucleotides can be determined even when a specific antibody or activity assay method is not available. These approaches will be useful for verifying the physiological role of other oncoproteins or proteins in living cells, and antisense oligonucleotides may be developed as new therapeutic agents.
Ethical and Legal Issues in the Procurement, Storage and Use of DNA
Authors: McQueen Matthew J.
Published: October 2004

At the risk of ignoring the benefits, the problems of genetic testing have been widely discussed. Many clinical laboratories are archival stores for human tissues and very few have formal policies and procedures for the use of this material. There is little evidence of planning or preparation of appropriate protocols for the safekeeping of the collected clinical material. The process of informed consent needs to be re-examined. The standard of what would be expected by a reasonable and prudent person poses challenging questions relating to confidentiality and privacy, control and ownership of the stored tissue and cells, withdrawing from a research study, length of storage of samples, the use of biological materials by other parties, and the use of biological materials for purposes other than those for which they were first obtained. There also needs to be a clear understanding of identifiable, non-identifiable and anonymous samples. Clinical laboratory staff and their professional organisations need to engage in the discussion and development around the ethical, legal and social issues raised by testing human DNA. Clinical research laboratories will be increasingly involved in DNA testing and will be asked for access to stored blood or tissue. It is essential that professional responsibility is understood and is then exercised in the presence of appropriate policies and procedures. There is also a paramount need to involve the public who are already demonstrating evidence of alienation from the world occupied by genetics research.
DNA Extraction and Stability for Epidemiological Studies
Authors: Visvikis Sophie , Schlenck Alexandra , Maurice Mickaël
Published: October 2004

During the last few years, an important development of molecular biology techniques has been observed in research and clinical laboratories, and consequently the availability of DNA is becoming essential for epidemiological studies. Several DNA extraction procedures have been proposed. However as the quantity and quality of the DNA extracted is very variable, standardization of the storage of samples, and of extraction procedures becomes essential.

Three steps will be considered. (i) Procedures of whole blood preservation prior to extraction which seem to affect yield and purity of the DNA extracted; (ii) DNA extraction procedures, which must be validated by a systematic DNA quality control using DNA molecular weight screening and spectrophotometric analysis, and also by verification that the DNA obtained is well adapted for molecular biology applications; (iii) the third important factor to study is the storage of DNA, since data on short-term (several months) and especially on long-term (several years) DNA stability is often missing and/or conflicting.

Detailed studies must be done in order to establish guidelines for proper handling of genetic material.
A Multilocus Genotyping Assay for Cardiovascular Disease
Authors: Cheng Suzanne , Pallaud Céline , Grow Michael A. , Scharf Stephen J. , Erlich Henry A. , Klitz William , Pullinger Clive R. , Malloy Mary J. , Kane John P. , Siest Gérard , Visvikis Sophie
Published: October 2004

In our efforts to develop diagnostic tests for complex multifactorial disorders, and to assist the research community in evaluating genetic markers for predisposition to cardiovascular disease, we have developed a prototype assay to genotype up to 35 variable sites among 15 genes. The candidate markers in this panel were selected from biological pathways likely to contribute to the development and progression of cardiovascular disease. Each sample is amplified in two multiplex polymerase chain reactions that are then hybridized to an array of immobilized oligonucleotide probes. The assay has been applied to a population-based cohort representing 238 families; allele frequencies observed among 455 unrelated parents from this cohort agree with available literature values. Data from a cohort of 142 lipid-clinic patients were used to explore locus associations with arterial occlusion, as measured by quantitative angiography. This prototype assay provides a research tool for studies to assess the association of multiple markers with disease, and for clinical studies to evaluate marker association with patient responsiveness to experimental therapies.
Quantification of Relative Expression of Genes with Homologous Sequences Using Fluorescence-Based Single-Strand Conformation Polymorphism Analysis – Application to Lactate Dehydrogenase and Cyclooxygenase Isozymes
Authors: Maekawa Masato , Sugano Kokichi
Published: October 2004

The combination of reverse transcribed-PCR and fluorescence-based single strand conformation polymorphism analysis has been proposed for the quantitative determination of ratio of mRNA molecules with homologous sequences. We applied this procedure to lactate dehydrogenase subunits M and H, and cyclooxygenase 1 and 2. We designed fluorescence labeled common PCR primers in the sequences highly homologous between two subunit and isozyme cDNAs, and performed reverse transcribed-PCR and fluorescencebased single strand conformation polymorphism analysis. PCR efficiency was almost the same for the different target sequences, so analysis of mixtures of known amounts of lactate dehydrogenase M and H revealed linear and precise proportions of lactate dehydrogenase M mRNA. It was shown that template concentrations and number of PCR cycles did not affect the determination of proportions of lactate dehydrogenase M to total lactate dehydrogenase. The procedure was applicable to a determination of cyclooxygenase-2 proportion; furthermore, the present procedure could be easily applied to investigation of expression levels of genes encoding mRNAs with homologous sequences.
A Simple PCR with Different 3′ Ends of the Third Primer for Detection of Defined Point Mutations: HCV Genotyping as an Example
Authors: Alavantić Dragan , Glisić Sanja , Radovanović Nataša , Romić Marija
Published: October 2004

In this article, we describe a useful modification of the polymerase chain reaction for amplification applicable to hepatitis C virus genotyping and determination of its subtypes. The method is fast, cheap and simple for detection of any known point mutation, and could be used in every laboratory with experience in polymerase chain reaction technique. We could differentiate hepatitis C virus subtype 1b from other subtypes and 2b from 2a and other subtypes as well. We could also differentiate hepatitis C type 3 using a type-specific oligonucleotide from 3a subtype, thus covering the most common hepatitis C virus (sub)types present in the European region.
Two Quantitative Methods for Imaging Samples in Molecular Biology
Authors: Tian Li
Published: October 2004

In this paper, we introduce two autoradiography imaging systems from Packard Instrument Company, that provide unmatched speed, accuracy, sensitivity, and resolving power for radioisotopic imaging in biomedical research. The first system is a direct nuclear imaging system, which provides fast and accurate nuclear counting without intermediate steps. Data are displayed in real time so that counting, imaging and data analysis can be done all at the same time. The second system is a new storage phosphor system that was designed with a unique optical assembly which provides high resolution imaging. With various types of phosphor screens, one can optimize the performance of this storage phosphor system with any radiolabel and any appropriate sample size. With these two systems, quantitative radioisotopic imaging becomes not only faster and easier, but also more consistent and accurate, providing researchers with a higher standard in molecular biology imaging.
Quantitative Detection of Hepatitis B Virus DNA with a New PCR Assay
Authors: Kessler Harald H. , Pierer Karen , Santner Brigitte I. , Vellimedu Srinivas K. , Stelzl Evelyn , Lackner Herwig , Moser Andrea , Marth Egon
Published: October 2004

The Amplicor™ HBV Monitor Test for quantitative detection of serum hepatitis B virus (HBV) DNA has recently been introduced. This assay is based on PCR and a non-radioactive hybridization and detection system on microwell plates. Evaluation in a routine diagnostic laboratory showed excellent sensitivity and adequate reproducibility; however, a more automated format would be desirable. The Amplicor™ HBV Monitor Test is useful for recognizing those patients who might benefit from antiviral treatment and for evaluation of the efficacy of anti-hepatitis B virus treatment.
Molecular Diagnostics for Cardiovascular Disease
Authors: Pang Chi-Pui
Published: October 2004

The etiology of cardiovascular diseases is known to be multi-factorial. Some forms of cardiovascular disease are influenced by unclear genetic factors but are predominantly affected by factors such as diet, obesity, cigarette smoking, diabetes mellitus and dyslipidaemia. Some are caused by specific gene defects, with environmental factors playing a precipitating role. Others result from complex gene-gene or gene-enviroment interactions. Advances in knowledge of the molecular genetics of lipidaemic and vascular disorders have identified gene aberrations that are associated with cardiovascular disease. Techniques in molecular biology have been applied for rapid and reliable detection of specific gene defects to provide unequivocal diagnosis beneficial for appropriate drug therapy and genetic counseling. Pre-symptomatic diagnosis is possible and carriers can be advised on effective preventive measures. However, prior to the provision of a molecular diagnostic service, all gene alterations associated with cardiovascular disease have to be identified and their prevalence established in a population. The number of mutations in so many causative genes is enormous. While more cost-effective laboratory methodologies will be developed in the future, it is also anticipated that more mutations with direct or indirect effects on cardiovascular disease will be discovered in different populations.
Association of Apolipoprotein E Polymorphism and Concentration with Serum Lipids and Apo-lipoprotein Level in the Chinese fromShanghai
Authors: Shen Xia , Xia Yongjing , Sass Catherine , Visvikis Sophie , Siest Gérard
Published: October 2004

The influence of apolipoprotein E polymorphism and apoE level on serum lipids and apolipoproteins was investigated in 71 healthy people and 43 patients with coronary artery disease from Shanghai. The frequency of apoE alleles was 0.06 for ε2, 0.86 for ε3, and 0.07 for ε4 in the healthy group, and 0.14 for ε2, 0.77 for ε3, and 0.09 for ε4 in the coronary artery disease group. There was no significant difference in the frequency of apoE alleles between these two groups. Serum levels of triglyceride and apo AI did not differ according to apoE genotypes, whereas serum level of apoB was significantly different according to apoE genotypes (p<0.05) both in healthy and coronary artery disease groups. However, in the healthy group, apo ε2 allele carriers had significantly higher level of apoE than apo ε3 and ε4 allele carriers (p<0.001) and apo ε4 allele carriers had significantly higher level of total cholesterol than apo ε3 and ε2 allele carriers. These were not observed in the coronary artery disease group. ApoE concentration was positively correlated with cholesterol, apoAI, and apoB levels in the control subjects and no significant correlation was observed with triglyceride level. In contrast, apoE level was positively related only to triglyceride level in the coronary artery disease group. In the control group, apoE genotypes and apoE level explained together 19.3 % and 26.6 % of the variability of apoB and cholesterol level, respectively, apoE polymorphism explained 23 % of the variability of apoE level and apoE level explained 13.2 % of the variability of apoAI level. In the coronary artery disease group, only apoE level explained 41.7 % of triglyceride variability. Finally we compared our results with those previously obtained in a French healthy population, the Stanislas cohort. Results suggested that there were some difference between the Chinese control and the French subjects.
Cholesteryl Ester Transfer Protein Mutations, Protein Activity and HDL-Cholesterol Concentration
Authors: Hill Stephen A. , Thomson Christopher , McQueen Matthew J.
Published: October 2004

We have examined a group of North American subjects, selected to include individuals with a wide variety of HDL-cholesterol concentrations for: 1) mutations in the genes coding for cholesteryl ester transfer protein and hepatic lipase, 2) apolipoprotein E genotype, 3) total cholesterol and triglycerides, 4) HDL-triglycerides. Cholesteryl ester transfer protein activity was also estimated, using a novel technique that does not require separation of substrate and product. Transfer activity was shown to have a monophasic distribution, with a mean activity of 21 pmol substrate transferred/3 h/μl plasma. The cholesterol ester transfer activity of the group with HDL-cholesterol >1.60 mmol/l was significantly less than those with HDL-cholesterol <1.60 mmol/l. The cholesteryl ester transfer protein G1533A mutation was detected at an overall allele frequency of 2.91 %. The mutation was more frequent in the group with HDL-cholesterol <1.60 mmol/l than in those >1.60 mmol/l. It was also more frequent in those with protein activity > 30 pmol/3h/μl plasma than in those with activity <30. These data suggest that this mutation in cholesteryl ester transfer protein is associated with increased transfer activity and reduced HDL-cholesterol concentrations. The cholesteryl ester transfer protein activity assay described here is simple and convenient. Subject to further evaluation and correlation with the present labour and time intensive assays, this commercially available assay offers the potential of rapid, simple analysis of large numbers of samples.
Molecular Biology of Alcohol Dependence, a Complex Polygenic Disorder
Authors: Whitfield John B. , Nightingale Brian N. , OBrien Martin E. , Heath Andrew C. , Birley Andrew J. , Martin Nicholas G.
Published: October 2004

Alcohol dependence, and the medical conditions which arise from prolonged excessive alcohol use, have no single cause. Like other complex diseases, they result from a combination of social, personal and genetic contributions; but within any society genetic variation has a substantial influence on individual risk. The genes presently known to affect alcohol dependence produce variation in alcohol metabolism; other genes which affect personality or susceptibility to intoxication are likely to be significant but so far reproducible evidence is scanty. Designs which include related subjects have advantages for the study of complex diseases, because any association effects can be placed in the context of overall heritability and because linkage analysis can also be included. Examples of our studies of alcohol metabolism, consumption and dependence are presented.
Screening for Tumour Suppressor p16(CDKN2A) Germline Mutations in Israeli Melanoma Families
Authors: Yakobson Emanuel A. , Zlotogorski Avraham , Shafir Rafael , Cohen Meir , Icekson Michael , Landau Marina , Brenner Sarah , Usher Sally , Peretz Hava
Published: October 2004

Approximately ten percent of patients with malignant melanoma have family histories of the disease, suggesting a genetic predisposition. Germline mutations in tumour suppressor p16 gene have been implicated as disease causing mutations in some of the melanoma families. The frequency of families with p16 germline mutations among melanoma prone families varies from eight to fifty percent. The range of the variability is influenced apparently by the number of melanoma affected individuals within the family, as well as by other, yet unidentified factors. Ethnic background is known to determine both the frequency and the nature of germline alterations. Recently, specific mutations in tumour suppressor genes involved in breast cancer and in colon cancer were found at elevated frequency among Ashkenazi Jews.

This report describes results of a screening for p16 germline alterations in a collection of Israeli melanoma families. We have analyzed genomic DNA from thirty one Ashkenazi and non-Ashkenazi Jewish melanoma families, as well as from thirty melanoma patients without an apparent family history of the disease. The entire coding region of the p16 gene was screened by single strand conformation polymorphism analysis and direct DNA sequencing.

We have detected a number of carriers with the Ala148 Thr polymorphism at the end of the second exon and several instances of 500(G⇒C) substitution at the 3′ untranslated portion of the gene.
Detection of CYP2D6*3 and 2D6*4 Allelic Variants by PCR-Restriction Fragment Length Polymorphism
Authors: Topić Elizabeta , Štefanović Mario , Nikolić Vjeran , Zoričić Ivan , Ivanišević Ana-Maria , Žuntar Irena
Published: October 2004

The mutant of CYP2D6*3 allele with A2637 deletion in exon 5 and the mutant of CYP2D6*4 allele G1934→A, splice site defect are among the most common polymorphic alleles of CYP2D6 gene, resulting in a decreased or no activity of CYP isoenzyme. In this study, a reliable polymerase chain reaction-restriction fragment length polymorphism method for identification of CYP2D6*3 and CYP2D6*4 alleles was used to investigate the genotype and phenotype prevalence in the groups of normal controls, and of cirrhosis and cancer patients. The results showed none of 36 controls genotyped for 2D6*3 and 2D6*4 allele to have the 2D6*3 allele with frameshift mutation in exon 5, while 33 % (n=12) were found to bear the 2D6*4 allele with G to A mutation at the intron 3—exon 4 junction. In breast cancer patients (n=35) genotyped for 2D6*3 and 2D6*4 alleles, none with 2D6*3 allele was found either, but 60 % (n=18) were found to bear the 2D6*4 allele. In patients with head and neck squamous cell cancer, there was only one subject with 2D6*3 allele and he was heterozygous. Among them, as many as ten (40 %) patients were found to bear 2D6*4 allele. In the cirrhosis group, none of the patients was found to have the 2D6*3 allele, while the CYP2D6*4 allele was found in 23 % (n=6) patients. The phenotype predicted according to the genotype was as follows: in the control group, 3% of individuals were identified as poor metabolizers, 70 % as extensive metabolizers, and 27 % as heterozygote extensive metabolizers. In the group of breast cancer, 7% of the patients were identified as poor metabolizer, 57 % as extensive metabolizer and 36% as phenotype. In squamous cell cancer and cirrhosis patients, the incidence of poor metabolizer was zero, and of heterozygotes extensive metabolizer 42 % and 31 %, respectively.
Follitropin (FSH) Deficiency in an Infertile Male due to FSHβ Gene Mutation. A Syndrome of Normal Puberty and Virilization but Under-developed Testicles with Azoospermia, Low FSH but High Lutropin and Normal Serum Testosterone Concentrations
Authors: Lindstedt Göran , Nyström Ernst , Matthews Clare , Ernest Ingrid , Janson Per Olof , Chatterjee Krishna
Published: October 2004

We studied a man who sought medical attention at age 28 years because of infertility in both his first and second marriages. His sexual development appeared to have been normal, with normal puberty and virilization, and normal libido and sexual potency. At examination, his testicles were small and soft; otherwise he had a normal physical appearance. Evaluations revealed azoospermia, undetectable in serum before and after 100 μg of intravenously administered gonadotrophin releasing hormone, but moderately elevated lutropin concentration with a brisk rise after gonadotrophin releasing hormone. The α subunit concentration was normal before and after gonadotrophin releasing hormone; that of inhibin B was very low. Analysis of the follitropin β gene, exon 3, revealed a Cys82 → Arg mutation (TGT → CGT). Judging from studies of the biosynthesis of the chorionic gonadotrophin β subunit one may conclude that inability to form the first intramolecular disulphide bond in the follitropin β subunit results in an abnormal tertiary structure during follitropin β biosynthesis with extensive intracellular degradation of the products, inability to associate with the α subunit and defective glycosylation, as well as inability to form a biologically active hormone. This first male case of follitropin deficiency thus defines a new syndrome of male infertility.
Molecular Analysis and Electromyoneurographic Abnormalities in Croatian Children with Proximal Spinal Muscular Atrophies
Authors: Barišić Nina , Sertić Jadranka , Billi Christopher , Barić Ivo , Sarnavka Vladimir , Babić Tomislav , Hrabač Pero , Begović Davor , Florentin Lina , Stavljenić-Rukavina Ana
Published: October 2004

Childhood onset proximal spinal muscular atrophy presents with considerable clinical variability. This study included 14 Croatian children aged 11 days to 8 years with spinal muscular atrophy types I-III verified clinically and electromyoneurographically. DNA of affected children was screened for deletions of exons 7 and 8 of the survival motor neuron gene and for deletion of exon 5 of the neuronal apoptosis inhibitor protein gene. Motor nerve conduction velocity and compound muscle action potential amplitude were decreased in children with spinal muscular atrophy type I and II. Deletions of exons 7 and 8 of the survival motor neuron gene and of exon 5 of the neuronal apoptosis inhibitor protein gene in children with spinal muscular atrophy type I-II suggested existence of more genetic abnormalities as compared to type III.

A decrease in compound muscle action potential amplitude and motor nerve conduction velocity in children with spinal muscular atrophy correlated with the disease severity, probably as a result of axonal degeneration.

Phenotypic severity in children onset spinal muscular atrophy is directly correlated with the extent of survival motor neuron and neuronal apoptosis inhibitor protein exon deletions.
Improved Laboratory Test Selection and Enhanced Perception of Test Results as Tools for Cost-Effective Medicine
Authors: Mayer Michael , Wilkinson Ian , Heikkinen Risto , Ørntoft Torben , Magid Erik
Published: October 2004

Inconsistencies in the way physicians perceive and handle identical laboratory results have untoward effects on morbidity, mortality and cost of medical care. In this context, the selection of suitable tests to answer definite clinical questions, and the manner in which laboratory results are presented have great impact on the action taken by the clinician.

This review addresses preferred methods to improve laboratory test selection, and examines methods that more effectively convey laboratory results to clinicians. It is anticipated that refined selection of tests, and presentation of the test results in a configuration that is easily perceived by the clinician, will facilitate interpretation of laboratory reports. Furthermore, any measures that promote the application of laboratory information in medical practice improve economics at the laboratory-clinical interface.

The presently described methods to optimize test selection and interpretation are:

likelihood ratios to provide estimates of the ability of a test to identify a clinical condition;

consensus-and discriminant function-analysis to estimate the performance of tests in diagnosing a particular disease or condition;

receiver operating characteristic (ROC) curves to assess discrimination capabilities.

The methods which improve test result perception are expression of results as multiples of the upper normal limit, utilizing signal strength to provide prognostic probabilities, and presentation of results in graphic forms that display mutually interrelated functions, with a specific cluster of results being highly suggestive of a given condition.

In addition, we discuss application of expert systems to provide rules based on knowledge and experience to analyze results of tests and suggest diagnosis and action, including additional tests when required.

It is anticipated that judicious utilization of laboratory services by application of the reviewed methodologies will help to achieve medically justified responses at a lower cost and help to achieve a proper balance between cost of tests and their clinical usefulness.
Effect of Haptoglobin Phenotypes on Growth ofStreptococcus pyogenes
Authors: Delanghe Joris , Langlois Michel , Ouyang Jin , Claeys Geert , Buyzere Marc De , Wuyts Birgitte
Published: October 2004

The haptoglobin (Hp) 2–1 and 2–2 phenotypes have been shown to agglutinateStreptococcus pyogenescarrying the membrane antigen T4. In this study, the growth rate of two strains ofStreptococcus pyogenes(T1 and T4) in human serum was compared among haptoglobin phenotypesin vitro. During incubation for 16 hours in serum of different haptoglobin types, only Hp 2–1 and Hp 2–2 sera showed an inhibitory effect on growth, Hp 2–2 being 1.85 times more potent than Hp 2–1. Growth ofStreptococcus pyogenesT4 negatively correlated with the serum concentration of Hp 2–1 (r = −0.908) and Hp 2–2 (r = −0.953). Haptoglobin-depleted serum had no inhibitory effect on bacterial growth. Addition of haemoglobin and ferric citrate to the serum accelerated the growth ofStreptococcus pyogenesT4 (P < 0.05) but not in Hp 2–2 serum. Haptoglobin types 2–1 and 2–2 can be regarded as inhibitors ofStreptococcus pyogenesgrowthin vitro. These data point towards a potential protective role of Hp 2–2 inStreptococcus pyogenesinfectionin vivo, independently of iron uptake.
The Cleavage of Pro-Urokinase Type Plasminogen Activator by Stromelysin-1
Authors: Orgel Dagmar , Schröder Werner , Hecker-Kia Adelheid , Weithmann Klaus Ulrich , Kolkenbrock Hansjörg , Ulbrich Norbert
Published: October 2004

Membrane binding of urokinase type plasminogen activator (u-PA) is thought to play a pivotal role in connective tissue remodeling and invasive processes. We compare the ability of different matrix-metalloproteinases involved in connective tissue turnover to cleave pro-urokinase type plasminogen activator between the catalytic domain and the receptor binding part to investigate a potential role for matrix-metalloproteinases in the regulation of membrane-associated proteolytic activity. We employed several forms of human stromelysin-1 (full length, C-truncated, and recombinant catalytic domain), rabbit C-truncated stromelysin-1, the human gelatinases A and B and the human catalytic domain of neutrophil collagenase. The gelatinases and the collagenase did not separate the receptor binding domain of pro-urokinase type plasminogen activator from the catalytic domain, whereas all stromelysin-1 forms cleaved the glutamic acid 143-leucine 144 bond of pro-urokinase type plasminogen activator. This reaction could be inhibited by specific inhibitors of matrix metalloproteinases and was not affected by inhibitors of serine proteinases. TheMr31000 cleavage product with leucine 144 as N-terminus displayed no proteolytic activity towards the prourokinase type plasminogen activator substrate pyro-Glu-Gly-Arg-pNA-HCl (S2444), but it could be activated by an additional treatment with plasmin. Comparison between full length stromelysin-1 and its C-truncated forms, showed that both exhibited the same cleavage properties towards pro-urokinase type plasminogen activator. Thus, the cleavage of pro-urokinase type plasminogen activator by stromelysin-1 is not influenced by the presence or absence of the C-terminal domain. The recombinant catalytic domain of MMP-3 generated pro-urokinase type plasminogen activator, whereas incubation of pro-urokinase type plasminogen activator with the native forms of human or rabbit stromelysin-1 led to a moderate activation of pro-uPA due to an additional cleavage that is catalyzed by a serine proteinase.
Cathepsin B in Human Leukocytes
Authors: Den Tandt Wilfried Richard , Scharpé Simon
Published: October 2004

We have applied a method using the fluorigenic substrate benzoloxycarbonyl-Arg-Arg-amido-4-methylcoumarin to measure cathepsin B, a thiol proteinase, in homogenates of human leukocytes. Data like pH optimum, stability, influence of thiol groups and effects of thiol proteinase inhibitors, lack of binding to Concanavalin A and lack of contribution to the fluorescence by other cathepsins indicate that cathepsin B is the enzyme masured.

Although the activity of the enzyme was linear with respect to time at all protein concentrations measured, there was an acceptable 10% deviation of the enzyme activity from linearity as a function of protein concentration. The enzyme in the homogenate was stable at 0 °C but was rapidly inactivated at 50 °C and above pH 6.5–7. Very limited activation on the one hand and variable inhibition on the other was seen by reagents containing thiol groups and thiol proteinase inhibitors respectively.

Latency (60% of the enzyme activity) indicates a probable subcellular lysosomal localization. There is no affinity towards the lectin Concanavalin A and the Km value was around 1 mmol/l. Normal enzyme activity values in leukocyte homogenates were determined.
Pattern of Plasma Cyclic Nucleotides and Related Hormones in Liver Cirrhosis and Hepatocellular Cacinoma
Authors: Bamonti-Catena Fabrizia , Peracchi Maddalena , Conte Dario , Cantalamessa Luigi , Lorenzano Elettra , Orsatti Alessandra , Salerno Francesco
Published: October 2004

To evaluate the pattern of plasma cyclic adenosine 3′,5‵-monophosphate, cyclic guanosine 3‵,5‵-monophosphate, atrial natriuretic factor and glucagon levels in different stages of chronic liver diseases, we measured these variables in 20 normal subjects, 25 patients with genetic hemochromatosis, associated with liver cirrhosis in 19 cases and not in six, eight patients with compensated and 15 with decompensated alcoholic or posthepatitic cirrhosis, and 12 with hepatocellular carcinoma. All variables were within the normal range in non-cirrhotic hemochromatotic patients. Cyclic adenosine 3‵,5‵-monophosphate levels were within the normal range (9.5–15.7 nmol/l) in hemochromatotic cirrhotics and elevated in other patients. Cyclic guanosine 3‵,5‵-monophosphate, atrial natriuretic factor and glucagon were above the normal ranges (1.92–5.91 nmol/l, 8.8–62.7 ng/l, and 39–165 ng/l, respectively) in most patients with cirrhosis both with and without hemochromatosis and in most individuals with hepatocellular carcinoma. Cyclic guanosine 3‵,5‵-monophosphate correlated with atrial natriuretic factor in the former groups but not in the latter. These findings indicate that glucagon and atrial natriuretic factor hypersecretion is an early event in cirrhosis, regardless of its etiology. In hepatocellular carcinoma, the underlying cirrhosis may account for most hormonal and metabolic changes although cyclic guanosine 3‵,5‵-monophosphate increases could also be due to the neoplastic process per se.
Serum Ionized versus Total Magnesium in Patients with Intestinal or Liver Disease
Authors: Saha Heikki , Harmoinen Aimo , Karvonen Anna-Liisa , Mustonen Jukka , Pasternack Amos
Published: October 2004

In serum, magnesium exists in three fractions: protein-bound, complex-bound and free ionized form. Only the free ionized fraction is biologically active. Until recently, only the measurement of serum total magnesium has been in clinical use. Now, commercially available instruments using new ion-selective electrodes for Mg++ have made possible the reliable measurement of serum ionized magnesium in clinical practice. For the measurement of serum ionized magnesium we used a magnesium-selective electrode installed in a six-channel electrolyte analyzer. We compared the use of ionized versus total magnesium measurement in 52 patients with intestinal disease, 54 with liver disease, and in 75 healthy control subjects.

In the patients with alcoholic liver disease both serum ionized and total magnesium were lower, and in those with inflammatory bowel disease slightly higher than in control subjects. The correlation coefficient between serum ionized and total magnesium was r=0.87 (p<0.001) in the patients, and r=0.75 (p<0.001) in the controls. In the patient group the fraction of ionized magnesium in the total was negatively related to the serum albumin level (r=−0.41, p<0.001). Serum total magnesium was below the reference range in 30 out of 150 measurements, serum ionized magnesium in only 9 out of 150 measurements, respectively. Thus, 21 cases with low total but normal ionized magnesium (two thirds of hypomagnesemia according to serum total magnesium) were false positive. Total magnesium measurement may overestimate the incidence of hypomagnesemia when significant hypoalbuminemia is present. Measurement of serum ionized magnesium instead of total magnesium may therefore be of advantage in evaluating patients with hypoalbuminemia and when hypomagnesemia is expected.
Evaluation of the Analytical Performances of a New Fully Automated Commercial Immunonephelometric Assay for Lipoprotein(a)
Authors: Lippi Giuseppe , Ruzzenente Orazio , Brentegani Claudio , Pierotti Anna , Guidi Giancesare
Published: October 2004

Several commercial methods have been proposed for lipoprotein(a) (Lp(a)) measurements over the past decades. However, only a few of them appear completely suitable in terms of analytical performance, costs and practicability. We evaluated the analytical performance of a new commercial fully automated immunonephelometric assay for Lp(a) measurements on the IMMAGE® Immunochemistry System. Mean within- and between-run coefficients of variation were 2.7 % (range 1.2–4.7 %) and 3.8 % (range 1.8–7.9 %), respectively. The linearity of the assay was confirmed up to 102 mg/dl and the deviation from the expected values did not exceed 4% (mean deviation: 1.9%). Moreover, the relative non-linearity was acceptable, ranging from 1.4 % to 1.6 % and hence constantly lower than the 2.5 % upper limit. Since there is no reference method for Lp(a) measurements, 100 routine random serum samples measured by the IMMAGE® Immunochemistry System immunonephelometric assay were further compared with two other commercial immunonephelometric assays (Array® LPA immunonephelometric assay and BNA Latex-Enhanced Lp(a) nephelometric assay). Non-parametric regression and relative Spearman's correlation coefficients were satisfactory, (y = 1.009x−1.38; r = 0.998 IMMAGE® Immunochemistry System vs. Array LPA and y = 0.922x−0.40; r = 0.989 IMMAGE® Immunochemistry System vs. Behring Nephelometer Analyzer (BNA) Latex Lp(a) assay). On the basis of the results of the present evaluation we conclude that the analytical performance and the main technical features of the IMMAGE® Immunochemistry System immunonephelometric assay make it a suitable method for Lp(a) measurement in clinical laboratories.
Immunoradiometric Assay of Thyroglobulin in Patients with Differentiated Thyroid Carcinomas: Need for Thyroglobulin Recovery Tests
Authors: Bourrel François , Hoff Madeleine , Regis Henri , Courrière Philippe , Caron Philippe
Published: October 2004

As interference from thyroglobulin autoantibodies appears to have been overcome in new commercial thyroglobulin assays by the use of monoclonal antibodies, the need for thyroglobulin recovery tests became uncertain. Sera (n=45) from patients with differentiated thyroid carcinomas were selected on the basis of a thyroglobulin recovery value below 70 % in the Dynotest Tg immunoradiometric assay (Brahms) routinely used in our laboratory. Serum thyroglobulin levels were then measured using three other commercial immunoradiometric assays: thyroglobulin ERIA (Pasteur), HTGK (Sorin) and ELSA HTG (Cis Bio International). Thyroglobulin autoantibodies were measured using the Thyrak assay (Brahms). Although many patients were thyroglobulin antibodies-negative (< 200 U/ml, n=26), most immunoradiometric assays failed to detect thyroglobulin in patients with evidence of recurrence. Low thyroglobulin values associated with low thyroglobulin recovery in thyroglobulin antibodynegative patients appear to be more biologically relevant than a single low thyroglobulin value, which can lead to lack of medical intervention. We conclude that the thyroglobulin recovery test is a prerequisite for the correct interpretation of serum thyroglobulin levels determined with immunoradiometric assays in the follow-up of thyroglobulin autoantibody-negative patients treated for differentiated thyroid carcinomas.
Effect of Thyroid Hormones on Lp(a) and Lipid Metabolism
Authors: Engler Hanna , Riesen Walter F.
Published: October 2004

The effect of thyroid hormones on lipoprotein(a) plasma concentrations and the other parameters of lipoprotein metabolism was studied in 158 patients with thyroid dysfunction and in 37 euthyroid controls (cross-sectional study). Multiple regression analysis revealed that 65.5 % of the variability in lipoprotein(a) levels were predicted by changes in lipoprotein(a) phenotypes (60.5 %), thyrotropin (3.5 %), and age (0.8 %). The lipid parameters, however, showed no significant effect on lipoprotein(a). A subgroup analysis on samples from patients with large lipoprotein(a) isoforms showed that 28% of the variability in lipoprotein(a) concentrations could be explained by changes in thyroid function (19.1 %), age (6.5 %) and triglycerides (3.5 %). Much stronger correlations were found between thyrotropin and total cholesterol, low density lipoprotein cholesterol or apolipoprotein B respectively (R2=0.951 for low density lipoprotein cholesterol, R2=0.801 for apolipoprotein B). Our data suggest that thyroid hormone is a significant modulator of lipoprotein(a) metabolism. However, different mechanisms are responsible for the change in lipoprotein(a) levels and for the decrease in total-and low density lipoprotein cholesterol levels.
Evaluation of the First Automated Thyroglobulin Assay
Authors: Vogeser Michael , Knesewitsch Peter , Jacob Karl , Seidel Dietrich
Published: October 2004

The aim of this study was to investigate technical and analytical performance of the first automated thyroglobulin (Tg) assay (DPC-Immulite®; Diagnostic Products Corporation, Los Angeles, USA). In imprecision studies using several human serum pools ranging from 21 to 58 replicates, a coefficient of variation of 9.0 % was obtained at a mean Tg concentration of 0.84 ng/ml and of 6.1 % at a Tg concentration of 62.1 ng/ml. In a method comparison with a non-automated assay (BRAHMS LUMItest Tg®, BRAHMS, Berlin, Germany) using 383 sera of 303 patients with thyroid carcinoma, regression analysis according to Passing and Bablock yielded in the following equation:

Immulite Tg=1.6 × BRAHMS Tg−0.1 ng/ml

(Pearson's r=0.979).

Sera obtained from 59 patients with thyroid carcinoma enabled comparative follow-up studies; in all cases qualitative agreement was found with regard to increase or decrease of serum Tg; in eight cases, however, Tg was detected with the Immulite assay but not with the BRAHMS assay. Further follow-up proved the presence of thyroid tissue in these patients. From these and further methodological data (dilution linearity, interference studies, carry-over study, high-dose hook properties, and short report time) it is concluded that the DPC-Immulite Tg assay meets the requirements of routine diagnostic use.
Lactoferrin: A Multifunctional Glycoprotein Involved in the Modulation of the Inflammatory Process
Authors: Baveye Sophie , Elass Elisabeth , Mazurier Joël , Spik Geneviève , Legrand Dominique
Published: October 2004

Lactoferrin is an iron-binding glycoprotein found in exocrine secretions of mammals and released from neutrophilic granules during inflammation. This review describes the biological roles of lactoferrin in host defence. Secreted lactoferrin exerts antimicrobial action either by chelation of iron or by destabilization of bacterial membranes. Furthermore, lactoferrin modulates the inflammatory process, mainly by preventing the release of cytokines from monocytes and by regulating the proliferation and differentiation of immune cells. Some of these activities are related to the ability of lactoferrin to bind lipopolysaccharides (LPS) with high affinity. Indeed, recentin vitrostudies indicate that lactoferrin is able to compete with the LPS-binding protein for LPS binding and therefore to prevent the transfer of LPS to CD14 present at the surface of monocytes. Moreover, the prophylactic properties of lactoferrin against septicemiain vivohave been demonstrated. Taken as a whole, these observations strongly suggest that lactoferrin is one of the key molecules which modulate the inflammatory response.