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Visual identification of individual transfected cells for electrophysiology using antibody-coated beads.

Electrophysiological study of transiently transfected cells requires the identification of individual cells that express the protein of interest. We describe a simple, quick and inexpensive method for visually identifying cells that have been co-transfected with an expression plasmid for a lymphocyte surface antigen (CD8-alpha). Transfected cells are incubated briefly with polystyrene microspheres (4.5 microns diameter) that have been precoated with antibody to CD8. Cells expressing CD8 on their surface are decorated with many beads and are thus readily distinguishable from untransfected cells. Beads already coated with antibody are available commercially. The method takes less than five minutes and requires no reagent preparation or special equipment for visualization of the beads.

Antibodies↗

[Identification of individuals with high coronary risk in the Italian population: indications of the Epidemiologic Cardiovascular Observatory].

BACKGROUND: The absolute global coronary risk has recently been introduced as an indicator of the incidence predicted by the main risk factors. It offers numerous options for the treatment of individuals at high risk. The identification of the absolute global coronary risk is produced through the application of functions obtained by longitudinal studies; their adequacy depends on the characteristics of the population from which they were estimated. The aim of this work was to evaluate the impact of the application of the absolute global coronary risk evaluation using the chart of risk proposed to the Italian physicians and to compare it with the results obtained from the application of other risk functions. METHODS: The database of the Osservatorio Epidemiologico Cardiovascolare (OEC), consisting of men and women aged 35-74 years, has been considered as being representative of the Italian population. The individual risk has been computed using the functions and coefficients from the Framingham study, the PROCAM study and the Seven Countries Study-Italy. The prevalence of high risk factors has been estimated on the basis of the recommendations on coronary prevention of the Task Force of the European Societies. RESULTS: The prevalence of high risk factors estimated by the Framingham function is 23.7% among men and 3.8% among women aged 35-74 years. In men aged 35-64 years, this estimated prevalence decreases from 14.2 to 8.7% when the Framingham function is adjusted using the mean value of the risk factors of the OEC, to 5.2% when the PROCAM function is applied, and to 1.1% when the function of the Seven Countries Study-Italy is employed. CONCLUSIONS: The application of the risk function suggested to the Italian physicians implies that more than 2,700,000 men and more than 500,000 women aged 35-74 years are potential candidates for treatment with lipid-lowering drugs. The comparison between the use of different functions in the OEC sample produces high numerical differences. The over-evaluation of the individual at high risk implies significant human and social costs. It is therefore essential to determine risk functions and coefficients derived from recent Italian studies including all age groups, both sexes and taking into account the different geographic characteristics of our country.

Adult↗

Migration from plasticized films into foods. 5. Identification of individual species in a polymeric plasticizer and their migration into foods.

To assess the significance of migration of polymeric plasticizers into foods, chemical characterization and quantification of individual oligomeric species is required. This paper reports the identification of seven individual oligomers isolated from a poly(butylene adipate) plasticizer. Based on mass spectrometry, NMR and chemical degradation, the oligomers were identified as a series of diol-terminated units ranging from a trimer up to an 11-monomer unit, along with a cyclic tetramer, all in the molecular weight range of 300-1100. A study of the migration of polymeric plasticizer from PVC film into olive oil indicated preferential migration of low molecular weight species. These oligomers which comprised 24% of the parent plasticizer contributed more than 90% of the plasticizer migration with the smallest oligomers migrating 90-fold more readily than the bulk of the plasticizer. From a knowledge of total polymeric plasticizer migration from PVC films under actual conditions of food-use, the abundance of individual oligomers in the foods has been estimated.

Adipates↗

Influence of HLA-DRB1 alleles on lymphoproliferative responses to a naturally processed and presented measles virus phosphoprotein in measles immunized individuals.

Identification of stimulatory T-cell epitopes recognized by CD4+ T lymphocytes is important for vaccine development. Our previous studies using mass spectrometry identified a naturally processed HLA class II restricted DRB1*0301 T cell epitope in the measles virus phosphoprotein, MV-P1 (residues 179-197). Here we provide lymphocyte proliferation data from peripheral blood mononuclear cells (PBMC) obtained from 131 HLA-DRB1*0301-positive and HLA-DRB1*0301-negative (HLA discordant) individuals previously immunized against measles and report that a single amino acid substitution in the MV-P1 T cell epitope can reduce T cell proliferation and CD4+ T-cell recognition. Measles virus and measles peptide-specific lymphoproliferative responses and HLA-DRB1 allele associations reveal that the DRB1*0701 allele provided suggestive evidence of association with both measles virus (p = 0.03) and MV-P1 peptide (p = 0.06) lymphoproliferation. A marginally significant increase in the frequency of the *0301 allele (p = 0.10) was found among subjects who demonstrated low cellular responses to the measles virus. We found no associations between proliferation levels to the MV-P1 and MV-P2 peptides with *0301 alleles. We speculate that the glutamic acid at position 192 of the measles phosphoprotein is a critical immunogenicity factor and may influence the antigenicity of the naturally processed HLA class II MV-P1 epitope.

Alleles↗

Soluble versus cell-bound CD4, CD8 from bronchoalveolar lavage: correlation with pulmonary diagnoses in human immunodeficiency virus-infected individuals.

Identification and assessment of cell populations in bronchoalveolar lavage (BAL) specimens may he used to follow the course of a disease state or response to specific therapy. Beyond cellular assessment, there are indications that the presence and quantity of soluble surface antigens released from activated cells may lead to improved understanding and facilitated diagnosis of a number of disease states. This study evaluated soluble markers (sCD4 and sCD8) in BAL and serum from HIV-infected individuals undergoing diagnostic bronchoscopy, and compared these values to flow cytometry-quantified BAL and peripheral blood cell CD4 and CD8. Patient pulmonary diagnosis (based on cytology and microbiology) was compared with patient blood and BAL-soluble and cell-bound CD4 and CD8 to determine the relationship of these markers to disease states in this population. Serum sCD8 in patients with fungal infections was significantly elevated above sCD8 in patients with Pneumocystis carinii or pulmonary bacterial infections, p = 0.0001. BAL sCD4/sCD8 ratio was also significantly different in patients with bacterial vs. fungal pulmonary infections, p = 0.01. These findings suggest that soluble markers, particularly elevated sCD8, may be an important indication of pulmonary disease progression in these HIV+ patients with fungal infections.

Bronchoalveolar Lavage Fluid↗

Identification of individuals by analysis of biallelic DNA markers, using PCR and solid-phase minisequencing.

We have developed a new method for forensic identification of individuals, in which a panel of biallelic DNA markers are amplified by the PCR, and the variable nucleotides are detected in the amplified DNA fragments by the solid-phase minisequencing method. A panel of 12 common polymorphic nucleotides located on different chromosomes with reported allele frequencies close to .5 were chosen for the test. The allele frequencies for most of the markers were found to be similar in the Finnish and other Caucasian populations. We also introduce a novel approach for rapid determination of the population frequencies of biallelic markers. By this approach we were able to determine the allele frequencies of the markers in the Finnish population, by quantitative analysis of three pooled DNA samples representing 3,000 individuals. The power of discrimination and exclusion of the solid-phase minisequencing typing test with 12 markers was similar to that of three VNTR markers that are routinely used in forensic analyses at our institute. The solid-phase minisequencing method was successfully applied to type paternity and forensic case samples. We also show that the quantitative nature of our method allows typing of mixed samples.

Adult↗

Fixed human tissues: a resource for the identification of individuals.

Polymorphic genetic loci of the deoxyribonucleic acid (DNA) present in formalin-fixed, paraffin-embedded tissues were successfully analyzed by utilizing the polymerase chain reaction. Using this analysis, with three different polymorphic loci [human leucocyte antigen (HLA) DQ alpha, low-density lipoprotein receptor, and parathyroid hormone], fixed tissues representing 14 different individuals were genotyped and could be distinguished from each other. The techniques were further applied to the fixed autopsy tissues of a man in which a question of paternity arose postmortem. Since many individuals have surgical procedures or autopsy, these readily available fixed tissues represent an additional resource for the identification of individuals.

Adult↗

Novel in vitro method for identification of individuals at risk for beryllium hypersensitivity.

Beryllium-specific lymphocytes were generated by in vitro immunization of peripheral blood mononuclear cells (PBMC) from healthy unexposed individuals. Measurement of blastogenic responses of PBMC by [3H]thymidine uptake demonstrated that sensitization of PBMC with beryllium salts followed by stimulation with unrelated salts resulted in a negative response, whereas sensitization and restimulation of PBMC with beryllium salts produced a positive response. Flow cytometric and cell depletion analyses showed that all of the responding cells were CD4+ T cells. The in vitro immunization system was used to screen 52 human subjects for susceptibility to beryllium sensitization in vitro. The results show that of the 52 healthy unexposed subjects tested, only 1 (2%) was highly responsive, 4 subjects (8%) were moderately responsive, 20 subjects (39%) were low-level responders, and 27 subjects (52%) were nonresponders. The results showing 2% high-level responsiveness to beryllium sensitization in vitro correlate with the 1 to 5% prevalence of chronic beryllium disease in individuals sensitized to beryllium dust in vivo and thus support the thesis that the in vitro immunization system may permit the identification of individuals at risk for beryllium hypersensitivity.

Adult↗

PLAC test for identification of individuals at increased risk for coronary heart disease.

Recent advances in cardiovascular research point to a critical role of inflammatory processes in the etiology of cardiovascular disease. This has led to the discovery of novel inflammatory biomarkers, which may be useful as additional screening tools for the identification of individuals at increased risk of coronary heart disease. One such novel inflammatory biomarker is lipoprotein-associated phospholipase A(2). This review discusses the recent development of a US Food and Drug Administration-approved blood test for lipoprotein-associated phospholipase A(2) (PLAC test, diaDexus, Inc.) and its efficacy as a predictive biomarker of risk for cardiovascular disease. More specifically, the article addresses the potential target group most likely to benefit from this new screening test and provides a prospective scenario for its implementation.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗

Coagulopathy and bleeding in the parturient patient. Recent information has helped in the identification of individuals at special risk.

Normal pregnancy is characterized by a hypercoaguable state that may progress to large or small vessel thrombosis and in some situations terminate in gross bleeding, due to consumption and lysis of clotting factors. Recent information concerning alterations in the factors that control normal clotting has helped in the identification of individuals at special risk for clotting and bleeding problems. At the present time, transfusion support is limited to unfractionated or semi-fractionated blood products. As knowledge is advanced, recombinant blood derivatives that will eliminate the risk of transmission of disease in the transfusion management of bleeding patients will emerge. In many patients consumption and lysis of clotting proteins occurs in catastrophic situations. In these situations the good clinical judgement of the obstetrician is of paramount importance. In order for the obstetrician to exercise this judgement most effectively, there must be good lines of communication among the clinicians involved in the patient's care and prompt, responsive support from the coagulation laboratory and the transfusion service.

Blood Coagulation Tests↗

Rapid, simple identification of individual osteoblastic cells and their specific products by cell blotting assay.

Biochemical and molecular biological studies of osteoblastic cell function and hormonal regulation are frequently confounded by the inherent cellular heterogeneity and phenotypic instability of existing in vitro and in vivo model systems. A new technique (derived from Western blotting or antibody-based detection of protein molecules bound to nitrocellulose paper) is described for identification of individual cells which synthesize osteoblast-specific gene products (bone Gla-protein, type I collagen, and alkaline phosphatase) or produce cAMP in response to parathyroid hormone (PTH) or isoproterenol. Dispersed primary neonatal rat calvariae or osteogenic sarcoma cells were "plated" on Immobilon-P (a hydrophobic transfer membrane with very high protein-binding capacity) for 30 minutes to several hours, followed by agonist treatment, formalin fixation, hematoxylin staining, and immunostaining with a battery of antibodies specific for osteoblastic products. Individual cells and their secretory zones were visualized by light microscopy and counted. Treatment with PTH with or without isoproterenol resulted in increases in the percentages of osteoblastic cells elaborating cAMP, as well as the intensity of immunostaining, but had no effects on MCF-7 cells, a nonosteoblastic breast carcinoma control line. The percentage of cells within each primary osteoblastic cell population isolated or rat osteogenic sarcoma cell clone (G2 or C12) that elaborated bone-specific proteins or that generated cAMP in response to PTH varied with time and the individual cellular preparation, reconfirming the cellular heterogeneity of these systems. This method, in conjunction with techniques such as in vitro hybridization, should prove useful in characterizing discrete osteoblastic bone cell subpopulations and in clarifying mechanisms of hormonal regulation by local and systemic agents.

Alkaline Phosphatase↗

The value of exercise-induced U-wave inversion on ECG chest wall mapping in the identification of individual coronary arterial lesions.

Exercise-induced U-wave inversion on chest wall mapping was compared with coronary arteriographic findings in 160 consecutive patients who presented with chest pain suggestive of ischaemic heart disease. ECG recordings were made from 16 points on the chest wall before, during and after exercise. None of the 27 patients with normal coronary arteriograms developed U-wave inversion during or after exercise (specificity = 100%). In 21 (all males) of the 133 patients (15.8%) with significant coronary arterial lesions, U-wave inversion on exercise was noticed on different coronary artery territories on the chest wall map, and its localization was correlated with angiographic evidence of individual coronary arterial lesions (100% projection rate). In 9 patients (6.8%) this sign was observed in the absence of any ST segment changes or Q waves. Exercise-induced U-wave inversion was the sole ECG criterion reflecting a lesion of the left anterior descending artery in 12 cases (9%), of the circumflex in 6 cases (4.5%), and in only one case of right coronary artery disease. This sign was not detectable in the conventional V5 site in 9 cases (7.1%) with significant disease of the left anterior descending coronary artery. These nine patients showed U-wave inversion on other areas of the left anterior descending coronary artery territory on exercise. Exercise-induced U-wave inversion disappeared in all the ten patients who underwent coronary artery bypass graft surgery. It is suggested that exercise-induced U-wave inversion shown on chest wall mapping is a reliable indicator of coronary artery disease, which disappears after myocardial revascularization, and in addition, aids identification of individual coronary arterial lesions.

Adult↗

Assessment of the WHO partial recording approach in identification of individuals highly susceptible to periodontitis.

The use of the WHO partial recording approach (CPITN) in the identification of 20-30-yr-old individuals with high past or present susceptibility to periodontitis was investigated in two studies in a factory population. Comparison with a traditional assessment in 149 subjects indicated that the probing depths and bleeding scores of the partial recording sites correlated well with the other sites which were less prone to periodontal disease than the CPITN sites, and that all the 23 subjects with deep pockets would have been identified by the use of the CPITN sites alone. However, when 75 of the subjects were re-examined using the partial recording sites, assessment of loss of clinical attachment levels indicated that only 16 of the 33 subjects who had loss of 2 mm or more clinical attachment at two or more sites had deep pockets, indicating that in this group assessment using pocket depth measurement alone seriously underestimated the proportion of individuals with a high past or present susceptibility to periodontitis.

Adult↗

Phase identification of individual crystalline particles by electron backscatter diffraction.

Recently, an electron backscatter diffraction (EBSD) system was developed that uses a 1024 x 1024 CCD camera coupled to a thin phosphor. This camera has been shown to produce excellent EBSD patterns. In this system, crystallographic information is determined from the EBSD pattern and coupled with the elemental information from energy or wavelength dispersive X-ray spectrometry. Identification of the crystalline phase of a sample is then made through a link to a commercial diffraction database. To date, this system has been applied almost exclusively to conventional, bulk samples that have been polished to a flat surface. In this investigation, we report on the application of the EBSD system to the phase identification analysis of individual micrometre and submicrometre particles rather than flat surfaces.

Journal Article↗

A historical perspective on the immunodiagnosis of schistosomiasis in China.

The investigation and application of immunodiagnosis in schistomiasis have had a history of about 50 years so far in China. The author, in this paper, briefly review the progress of the technology involved here in the country, putting the stress on the latest issues during last decade. Also showed some limits in the use for control program of the disease through summary of some collaborative studies with the attempt in identifying guidelines for the implementation of serology in schistosomiasis immunodiagnosis. In addition, it was pointed out that a seroepidemiological approach in the identification of target individuals for treatment or re-treatment may be realistic and practical, which emphasized that the immunodiagnostic approach for public health use could ignore individual identification of the real infections. In this way, it became possible to bypass the complexity of interpreting the seroreactivity of an individual in assigning him/her with qualitative testing to the normal or infected person. Instead, one needs only to divide individuals in a community into 'normal' or 'infected' groups according to the titer distribution, which leads to identifying target groups for treatment.

China↗

[Methods of making the plaster copies of skulls for the medical and criminalistic identification of individuals].

A possibility was studied to use the plaster copies of skulls in the forensic medical identification of personality by applying the craniofacial method. No dependence was found between the plaster casts and the moulding types. Differences between the same craniometric sizes in the skull and in its cast are of an incidental nature. Such incidental deviation is related with an accuracy of determining the localization of craniometric points and with an instrumental precision of measuring devices. The necessity is substantiated that, while making a craniofacial examination, it is necessary to mark all anthropometric points in the skull cast, with the original being a control. The use of a plaster skull cast is possible in those cases, when the burial of the remains must be done in full.

Calcium Sulfate↗

[The kinological identification of individual scents in the traces of the vital activities of 4 vertebrate species].

One can successfully identify individual odors of almost any terrestrial vertebrates using laboratory dogs. The excretions can be collected on adsorbent paper and conserved for subsequent identification. It has been experimentally shown that the odor of house mice remains stable during the whole life span and that the stabilization of dog-identifiable odors finishes in young mice at 14 day. The method tested will allow to create individual banks of animal odors, e.g. for identification of rare species.

Animals↗

Evaluation of the Anaerobe-Tek system for identification of anaerobic bacteria.

The Anaerobe-Tek system of Flow Laboratories, Inc. (McLean, Va.), is a new bacterial identification system which uses Gram morphology, sporulation, and reactions from 15 agar-based biochemical tests to generate a six-digit code number for identification of anaerobic bacteria. Supplemental tests are recommended when necessary to complete species identification. Individual test and identification performance of this system was evaluated by testing 216 anaerobic bacteria representing 31 species and one Centers for Disease Control unnamed group in parallel with a routine clinical laboratory identification system. Most of the tests in the Anaerobe-Tek system performed well; 85% of the organisms were correctly identified. The 32 (15%) failures in identification were due to omission from the identification code data base (38%), false-negative indole reactions (22%), and other incorrect biochemical reactions (40%). The replacement of the recommended indole test with an extraction method using the inoculum broth and an expansion of the data base of the system could raise the correct identification for this organism population to over 90% with no change in the test materials.

Anaerobiosis↗