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PubMed1990

Applicability of the popliteal lymph node assay in the Brown-Norway rat.

A number of drugs and chemicals can induce autoimmune disorders. Previous studies suggested the popliteal lymph node (PLN) assay might be a suitable and reliable model to detect such compounds in the mouse. The applicability of this assay has been examined in the Brown-Norway rat using a panel of positive as well as negative reference compounds. Popliteal lymph nodes were excised and weighed seven days after subcutaneous injection of 5 mg of each compound into one footpad. Interestingly, all negative reference compounds produced no significant increase in PLN weight index whereas four positive reference compounds were associated with PLN-weight indices greater than 2.3. However, no increase in PLN weight was noted in procainamide- and isoniazide-treated rats as previously reported in the mouse. This first set of experiments indicates that the PLN assay is applicable to Brown-Norway rats and warrants further investigation.

Animals↗

PubMed1999

A high-density integrated genetic linkage and radiation hybrid map of the laboratory rat.

The laboratory rat (Rattus norvegicus) is a key animal model for biomedical research. However, the genetic infrastructure required for connecting phenotype and genotype in the rat is currently incomplete. Here, we report the construction and integration of two genomic maps: a dense genetic linkage map of the rat and the first radiation hybrid (RH) map of the rat. The genetic map was constructed in two F2 intercrosses (SHRSP x BN and FHH x ACI), containing a total of 4736 simple sequence length polymorphism (SSLP) markers. Allele sizes for 4328 of the genetic markers were characterized in 48 of the most commonly used inbred strains. The RH map is a lod >/= 3 framework map, including 983 SSLPs, thereby allowing integration with markers on various genetic maps and with markers mapped on the RH panel. Together, the maps provide an integrated reference to >3000 genes and ESTs and >8500 genetic markers (5211 of our SSLPs and >3500 SSLPs developed by other groups). [Bihoreau et al. (1997); James and Tanigami, RHdb (http:www.ebi.ac.uk/RHdb/index.html); Wilder (http://www.nih.gov/niams/scientific/ratgbase); Serikawa et al. (1992); RATMAP server (http://ratmap.gen.gu.se)] RH maps (v. 2.0) have been posted on our web sites at http://goliath.ifrc.mcw.edu/LGR/index.html or http://curatools.curagen.com/ratmap. Both web sites provide an RH mapping server where investigators can localize their own RH vectors relative to this map. The raw data have been deposited in the RHdb database. Taken together, these maps provide the basic tools for rat genomics. The RH map provides the means to rapidly localize genetic markers, genes, and ESTs within the rat genome. These maps provide the basic tools for rat genomics. They will facilitate studies of multifactorial disease and functional genomics, allow construction of physical maps, and provide a scaffold for both directed and large-scale sequencing efforts and comparative genomics in this important experimental organism.

Alleles↗

PubMed2002

Limits of diagnostic accuracy of anti-hepatitis C virus antibodies detection by ELISA and immunoblot assay.

When human sera samples are tested for anti-hepatitis C virus (HCV) antibodies using different ELISA kits as well as immunoblot assay kits discrepant results often occur. As a result the diagnostics of HCV infection in such sera remains unclear. The purpose of this investigation is to define the limits of HCV serodiagnostics. Overall 7 different test kits of domestic and foreign manufacturers were used for the sampled sera testing. Preliminary comparative study, using seroconversion panels PHV905, PHV907, PHV908 was performed and reference kit was chosen (Murex anti-HCV version 4) as the most sensitive kit on the base of this study results. Overall 1640 sera samples have been screened using different anti-HCV ELISA kits and 667 of them gave discrepant results in at least two kits. These sera were then tested using three anti-HCV ELISA kits (first set of 377 samples) or four anti-HCV ELISA kits (second set of 290 samples) at the conditions of reference laboratory. In the first set 17.2% samples remained discrepant and in the second set - 13.4%. "Discrepant" sera were further tested in RIBA 3.0 and INNO-LIA immunoblot confirmatory assays, but approximately 5-7% of them remained undetermined after all the tests. For the samples with signal-to-cutoff ratio higher than 3.0 high rate of result consistency by reference, ELISA routing and INNO-LIA immunoblot assay was observed. On the other hand the results of tests 27 "problematic" sera in RIBA 3.0 and INNO-LIA were consistent only in 55.5% cases. Analysis of the antigen spectrum reactive with antibodies in "problematic" sera, demonstrated predominance of Core, NS3 and NS4 antigens for sera, positive in RIBA 3.0 and Core and NS3 antigens for sera, positive in INNO-LIA. To overcome the problem of undetermined sera, methods based on other principles, as well as alternative criteria of HCV infection diagnostics are discussed.

Enzyme-Linked Immunosorbent Assay↗

PubMed1991

A simplified and less expensive strategy for confirming anti HIV-1 screening results in a diagnostic laboratory in Lubumbashi, Zaire.

The conventional algorithm for HIV testing based on the confirmation of all positive anti-HIV screening reactions by Western blot (WB) is too expensive for developing countries. We investigated the validity of confirming positive screening assay reactions by a second screening test, limiting the use of the supplemental assay to the discrepant test results (algorithm 3), or screening all sera with 2 different assays and retesting all discrepant results by a supplemental assay (algorithm 4) on a panel of 519 sera in a regional reference laboratory in Lubumbashi, Zaire. Combining the Vironostika anti-HTLV-III ELISA with HIV Chek 1 + 2 or Clonatec Rapid HIV 1/2 Ab on all samples and retesting the discrepant results in WB or a line immunoassay (INNO-LIA) (algorithm 4), yielded a sensitivity of 100% and specificities of 98.4% and 99.0% respectively, at costs of 7.3 US $ and 9.3 US $ per test, respectively, for a 40% prevalence of HIV antibody positive samples. The conventional algorithm scored a sensitivity of 97.1% and a specificity of 100% for 11.3 US $ per test. The testing strategy of combining HIV Chek 1 + 2 and Clonatec Rapid HIV 1/2 Ab, an interesting option for small isolated centra, had a 96.6% sensitivity, but yielded only a slightly better specificity of 99.0%, as compared to 97.8% for HIV Chek alone. The price of combining the two simple assays using algorithm 3 was 6.8 US $ per test, using algorithm 4 was 10.6 US $. HIV testing strategies based on ELISA and a simple HIV test are a valuable alternative for reference laboratories faced with a high prevalence of HIV positive samples.

AIDS Serodiagnosis↗

PubMed1990

Antigenic comparisons of hog cholera virus isolates from Europe, America and Asia using monoclonal antibodies.

Nineteen monoclonal antibodies (MAbs) with specificity for hog cholera virus (HCV) were prepared. They were used in an immune binding (peroxidase linked) assay to determine the reaction patterns of HCV isolates from Europe, Brazil, USA, Japan and Malaysia, as well as laboratory reference strains of the virus. A further panel of 17 MAbs raised against bovine virus diarrhoea virus (BVDV) was included in the study, together with 5 MAbs raised against a non-HCV pestivirus of porcine origin. All the MAbs were also tested against representative strains of BVDV and border disease virus. Six MAbs were HCV-specific, reacting with all isolates of HCV and none of the ruminant viruses. Among the other HCV MAbs geographical variation in reaction patterns was observed. There was evidence of antigenic distinction between recent European isolates, and archive material originally isolated more than 10 years ago.

Animals↗

PubMed1993

Report of the Canadian Hypertension Society Consensus Conference: 2. Diagnosis of hypertension in adults.

OBJECTIVE: To update recommendations for the diagnosis of mild hypertension in adults and to assess the role of echocardiography, self-measurement of blood pressure and ambulatory blood pressure monitoring. DATA SOURCES: Literature reviews of previous consensus conferences were updated with searches of MEDLINE for the period Jan. 1, 1988, to Nov. 15, 1991, and supplemented by reference lists and personal files. STUDY SELECTION: Panel members selected relevant articles and rated them according to methodologic criteria. DATA EXTRACTION: The data extracted concerned the measurement of blood pressure, the diagnosis of hypertension, the treatment of mild hypertension, and the reliability and validity of echocardiography, self-measurement of blood pressure and ambulatory blood pressure monitoring in the diagnosis of mild hypertension. The recommendations made were graded according to the level of evidence available, circulated to many experts and approved at a consensus conference. MAIN RESULTS: Previous recommendations for the accurate measurement of blood pressure remain mostly unchanged. Antihypertensive treatment should be prescribed for patients (including the elderly) with an average diastolic blood pressure of at least 100 mm Hg, for those with isolated systolic hypertension (systolic blood pressure of at least 160 mm Hg and diastolic blood pressure of less than 90 mm Hg) and for patients with a diastolic blood pressure of 90 to 99 mm Hg and target-organ damage. Clinical judgement is required in treating patients with a diastolic blood pressure of 90 to 99 mm Hg without target-organ damage, and individual risk for cardiovascular disease must be taken into account. There is insufficient evidence to warrant the routine use of echocardiography, self-measurement of blood pressure or ambulatory blood pressure monitoring in diagnosis. CONCLUSIONS: Recent high-quality evidence supports several new recommendations for the diagnosis of mild hypertension in adults. Additional research is needed to determine the role of echocardiography, self-measurement of blood pressure and ambulatory blood pressure monitoring.

Adult↗

PubMed1996

Osteoclast differentiation antigen.

Osteoclasts are the primary cells which perform bone resorption. The origin of these multinucleated giant cells is the haematopoietic stem cells. The differentiation pathway of the osteoclasts has so far been well studied and the cell-lineage of these bone resorbing cells is considered to be close but not identical to the monocytes/macrophages. Owing to the development of in vitro culture systems for evaluating osteoclast differentiation, it has been elucidated that various cytokines are involved in the differentiation of the osteoclasts. However, there is still ambiguity concerning the molecular mechanism of the differentiation of the osteoclasts. One approach for clarifying the molecular mechanism is to find unique antigen molecules involved in the process of osteoclast differentiation. In this review article, we introduce such immunological studies concerning osteoclast differentiation. We also refer to our recent establishment of a panel of monoclonal antibodies recognizing rat osteoclasts. One of the monoclonal antibodies recognizes cell surface antigen (Kat1-antigen) expressed on cells in osteoclast-lineage and not on monocytes/macrophages. Cross-linking of the cell surface antigen using this monoclonal antibody showed that the Kat1-antigen is the unique cell surface molecule involved in the regulation of the affinity of the calcitonin receptor and also involved in the modulation of bone resorption. In this review article, we overview, the current issues which should be elucidated for understanding the differentiation and activation of the osteoclasts. We further emphasize the utility of the immunological approach for solving these current target issues.

Animals↗

PubMedDate not supplied

Serotype identification of avian infectious bronchitis virus by RT-PCR of the peplomer (S-1) gene.

The S-1 peplomer gene sequences of 31 strains of avian coronavirus infectious bronchitis virus (IBV) from North America, Europe, and Australia were compared to identify common and unique regions for possible diagnostic applications. S-1 sequences that were conserved among serotypes and sequences that were variable between serotypes were identified. Based on conserved S-1 gene sequences, "general" degenerate oligonucleotide primers were designed that amplified IBV genomic RNA by the reverse transcriptase polymerase chain reaction (RT-PCR) procedure regardless of serotype. Primers specific for IBV serotypes Massachusetts, Connecticut, Arkansas, JMK, Delaware (DE/072/92), and California (CA/633/85) were designed from regions of the S-1 gene exhibiting extensive sequence hypervariability. The ability to identify these six serotypes of IBV by RT-PCR was demonstrated by testing the serotype-specific primers on a panel of unknown samples that included 30 reference strains and field isolates previously characterized by virus neutralization (VN). The use of serotype-specific primers in RT-PCR provides a rapid and accurate means of identifying IBV.

Amino Acid Sequence↗

PubMedDate not supplied

Comparison of susceptibilities of anaerobic bacteria determined by agar dilution and by a microbroth method.

The minimal inhibitory concentrations (MICs) of 110 anaerobic bacteria (84% fresh clinical isolates) to nine antimicrobial agents were determined simultaneously by the tentative reference agar-dilution method of the National Committee for Clinical Laboratory Standards and a prototype commercial microbroth panel (Microscan). MICs (determined visually for either system) differed by greater than 1 log2 dilution in 17% of all tests. With the exceptions of the MICs of tetracycline and moxalactam, however, all compared antimicrobial MICs were within 1 log2 dilution for at least 82% of all tests. Twenty-three anaerobic bacterial strains (21%) failed to grow in the microdilution panel (Wilkins-Chalgren broth), and seven strains (6%) failed to grow on the reference Wilkins-Chalgren agar. However, for those bacterial strains that could be tested, the microbroth panel appeared to be less cumbersome than the agar-dilution procedure for susceptibility testing of anaerobic microorganisms.

Agar↗

PubMed2001

Heteroduplex panel analysis, a novel method for genetic identification of Aspergillus Section Flavi strains.

For genetic identification of Aspergillus Section Flavi isolates and detection of intraspecific variation, we developed a novel method for heteroduplex panel analysis (HPA) utilizing fragments of the internal transcribed spacer (ITS) regions (ITS1-5.8S-ITS2) of the rRNA gene that was PCR amplified with universal primers. The method involves formation of heteroduplexes with a set of reference fragments amplified from Aspergillus flavus, A. parasiticus, A. tamarii, and A. nomius and subsequent minislab vinyl polymer gel electrophoresis. The test panel is compared with species-specific standard panels (F-1, P-1, T-1, and N-1) generated by pairwise reannealing among four reference fragments. Of 90 test panels, 89 succeeded in identifying the species and 74 were identical to one of the four standard panels. Of the 16 new panels, 11 A. flavus/A. oryzae panels were identical and typed as F-2 and 4 of 5 A. nomius panels were typed as N-2 or N-3. The other strain, A. nomius IMI 358749, was unable to identify the species because no single bands were formed with any of the four reference strains. DNA sequencing revealed that our HPA method has the highest sensitivity available and is able to detect as little as one nucleotide of diversity within the species. When Penicillium or non-Section Flavi Aspergillus was subjected to HPA, the resulting bands of heteroduplexes showed apparently lower mobility and poor heteroduplex formation. This indicates that HPA is a useful identification method without morphological observation and is suitable for rapid and inexpensive screening of large numbers of isolates. The HPA typing coincided with the taxonomy of Section Flavi and is therefore applicable as an alternative to the conventional methods (Samson, R. A., E. S. Hoekstra, J. C. Frisvad, and O. Filtenborg, p. 64-97, in Introduction to Food- and Airborne Fungi, 6th ed., 2000).

Aspergillus↗

PubMed2006

An alternate line erasure and readout (ALER) method for implementing slot-scan imaging technique with a flat-panel detector--initial experiences.

This paper describes and demonstrates an electronic collimation method, referred to as the alternate line erasure and readout (ALER) technique, for implementing slot-scan digital radiography technique with an amorphous silicon (a-Si) thin-film transistor (TFT) array based flat-panel detector. An amorphus selenium (a-Se) flat-panel detector was modified to implement the ALER technique for slot-scan imaging. A stepping-motor driven fore-collimator was mounted in front of an X-ray tube to generate a scanning X-ray fan beam. The scanning speed and magnification were adjusted to synchronize the fan beam motion with the image line readout rate. The image lines on the leading and trailing edges of the fan beam were tracked and alternately reset and read out, respectively. The former operation resulted in the erasure of the scatter signals accumulated in the leading edge image line prior to the arrival of the fan beam. The latter operation resulted in the acquisition of fan beam exposure data integrated in the trailing edge image line right after the fan beam passed. To demonstrate the scatter rejection capability of this technique, an anthropomorphic chest phantom was placed in PA position and scanned at a speed of 576 lines (8.0 cm)/s at 117 kVp and 32 mA. A tungsten bar is placed at the entrance side of the chest phantom to measure the scatter-to-primary ratio (SPR), scatter reduction factor (SRF), and contrast-to-noise ratio degradation factor (CNRDF) in the slot-scan images to evaluate the effectiveness of scatter rejection and the resultant improvement of image quality. SPR and CNRDF in the open-field images were also measured and used as the reference for comparison. A scatter reduction by 86.4 to 95.4% across lower lung and heart regions has been observed with slot-scan imaging. The CNRs have been found to be improved by a factor of 2 in the mediastinum areas over the open-field image as well.

Algorithms↗

PubMed1993

Numerical approach to reference identification of Staphylococcus, Stomatococcus, and Micrococcus spp.

A numerical-code system for the reference identification of Staphylococcus species, Stomatococcus mucilaginosus, and Micrococcus species was established by using a selected panel of conventional biochemicals. Results from 824 cultures (289 eye isolate cultures, 147 reference strains, and 388 known control strains) were used to generate a list of 354 identification code numbers. Each six-digit code number was based on results from 18 conventional biochemical reactions. Seven milliliters of purple agar base with 1% sterile carbohydrate solution added was poured into 60-mm-diameter agar plates. All biochemical tests were inoculated with 1 drop of a heavy broth suspension, incubated at 35 degrees C, and read daily for 3 days. All reactions were read and interpreted by the method of Kloos et al. (G. A. Hebert, C. G. Crowder, G. A. Hancock, W. R. Jarvis, and C. Thornsberry, J. Clin. Microbiol. 26:1939-1949, 1988; W. E. Kloos and D. W. Lambe, Jr., P. 222-237, in A. Balows, W. J. Hansler, Jr., K. L. Herrmann, H. D. Isenberg, and H. J. Shadomy, ed., Manual of Clinical Microbiology, 5th ed., 1991). This modified reference identification method was 96 to 98% accurate and could have value in reference and public health laboratory settings.

Bacterial Typing Techniques↗

PubMed1992

Clinicians' quick reference guide to acute pain management in infants, children, and adolescents: operative and medical procedures. Pain Management Guideline Panel. Agency for Health Care Policy and Research, US Department of Health and Human Services.

The management of pain is an important part of a health professional's role. The challenge for clinicians is to balance pain control with concern for patient safety and side effects of pain treatments. This is the second of two articles presenting the clinical highlights of the Agency for Health Care Policy and Research's Panel to develop clinical practice guidelines for pain management. This article presents the clinical highlights related to the assessment and management of pain in infants and children.

Acute Disease↗

PubMed1993

Specific detection of anti-HBc antibodies with an enzyme immunoassay using recombinant HBcAg and monoclonal antibodies.

An enzyme immunoassay for the detection of total anti-HBc antibodies in undiluted serum samples was developed. This assay utilizes an anti-HBc monoclonal antibody and a recombinant HBc antigen. The results of the clinical validation are now reported. A total of 1,301 sera were tested using both the Recombinant TOTAL HBc Ab EIA and a reference assay. The specificity was evaluated on a panel of 573 normal human sera and human sera from subjects with pathological findings unrelated to a hepatitis B virus infection. The sensitivity was studied on a total of 455 sera from HBV infected patients at different stages of infection. The final results indicate 99.8% sensitivity and 99.8% specificity. In addition, 273 sera with either isolated anti-HBc antibodies or with anti-HCV antibodies were tested. The agreement between the Recombinant and the reference assay for these two populations, 96.6 and 90.1%, respectively, is discussed.

Acute Disease↗

PubMed2006

Development of a framework for international certification by OIE of diagnostic tests validated as fit for purpose.

Historically, the OIE has focused on test methods applicable to trade and the international movement of animals and animal products. With its expanding role as the World Organisation for Animal Health, the OIE has recognised the need to evaluate test methods relative to specific diagnostic applications other than trade. In collaboration with its international partners, the OIE solicited input from experts through consultants' meetings on the development of guidelines for validation and certification of diagnostic assays for infectious animal diseases. Recommendations from the first meeting were formally adopted and have subsequently been acted upon by the OIE. A validation template has been developed that specifically requires a test to be fit or suited for its intended purpose (e.g. as a screening or a confirmatory test). This is a key criterion for validation. The template incorporates four distinct stages of validation, each of which has bearing on the evaluation of fitness for purpose. The OIE has just recently created a registry for diagnostic tests that fulfil these validation requirements. Assay developers are invited to submit validation dossiers to the OIE for evaluation by a panel of experts. Recognising that validation is an incremental process, tests methods achieving at least the first stages of validation may be provisionally accepted. To provide additional confidence in assay performance, the OIE, through its network of Reference Laboratories, has embarked on the development of evaluation panels. These panels would contain specially selected test samples that would assist in verifying fitness for purpose.

Animal Diseases↗

PubMed1985

HEP versus PNU standardization of allergen extracts in skin prick testing. A comparative randomized in vivo study.

During 1 year, all patients referred to an allergy outpatient clinic for adults were skin prick tested with a panel of standard allergen extracts from two manufacturers using different methods of standardization. One company referred to the histamine equivalent prick (HEP) and the other used the more traditional protein nitrogen units (PNU). Standard extracts and five-fold dilution were tested. The results indicate that the ratio of concentration between two extracts of the same allergen should be measured by the absolute difference of the wheal diameters. We found significant differences between corresponding extracts from the two manufacturers.

Adult↗

PubMed1992

Design and preliminary performance characteristics of a newly proposed reference cell for ionized calcium in serum.

A novel flow-through design for a calcium ion-selective liquid membrane and half-cell is proposed. The membrane is based on a neutral carrier for calcium and has the same quantitative chemical composition as that used in most commercial analyzers for ionized calcium (iCa2+). The membrane and half-cell follow the specifications in the Stage 1, Draft 4 document for a Reference Method for iCa2+ of the IFCC Expert Panel on pH, Blood Gases, and Electrolytes. Measurements of iCa2+ in serum with this cell confirm prior observations that protein adsorption on the membrane surface causes a rapid but reversible shift in the cell calibration signal and possible systematic errors in the measurement of iCa2+ in serum. When the protein adsorption phenomenon is maintained at equilibrium during calibrations and sample measurements, the new cell can be used to reproducibly assign iCa2+ concentrations to serum pools. These calibrated pools may then be used as reference materials and to resolve the biases known to exist among commercial iCa2+ analyzers.

Calcium↗

PubMed2003

[Quality control of the serological diagnosis of dengue in laboratories throughout the Americas, 1996-2001].

OBJECTIVE: To report the results from participating laboratories for four external quality control proficiency tests of dengue serological diagnosis that were carried out in the Region of the Americas in the period of 1996-2001. METHODS: External quality control proficiency tests of dengue serological diagnosis were carried out in 1996-1997, 1998-1999, 1999-2000, and 2000-2001. Panels made up of 20 serum samples (12 of them positive for dengue IgM antibodies) were sent to participating laboratories in the Region. The sera were negative for HIV antibodies, hepatitis C virus antibodies, and hepatitis B surface antigen. The sera were stored at -20 degrees C until they were sent in refrigerated shipments to the participating laboratories. The presence of IgM antibodies was determined through IgM-capture enzyme-linked immunosorbent assay (ELISA), while the IgG antibody titer was determined by hemagglutination inhibition or by IgG ELISA. The results of the IgM antibody testing that differed from those of the reference center were considered discordant. The IgG antibody titer was considered discordant when the results differed by two dilutions or more with respect to the reference center's results. RESULTS: A total of 27 laboratories received a total of 59 serum panels over the 1996-2001 period, and the results from testing 54 of those panels (91.5%) were sent back in. Of the total of 1 080 sera samples from those 54 panels, the results from 95.6% of the IgM antibody tests were concordant with the results from the reference center. With 47 of the 54 panels (87.0%) the participating laboratories' agreement with the reference center's results for the IgM antibody testing was 90.0% or higher. The laboratories sent back results from a total of 27 IgG antibody titer tests, and 22 of them (81.5%) coincided with those from the reference center. Considering the IgM antibody testing results from the four periods, the findings from 22 of the participating laboratories coincided with those from the reference center for at least 90% of the samples, and 13 of the laboratories were in complete concordance with the reference center. CONCLUSIONS: The majority of the participating laboratories showed an excellent level of performance in detecting dengue IgG and IgM antibodies. However, the deficiencies found in some instances confirm the need for continuing to improve laboratory diagnosis of dengue in the Region of the Americas.

Americas↗