Comparison of Antistreptolysin O Latex Screening Test with the Antistreptolysin O Hemolytic Test

This report describes a comparison of the Behringwerke antistreptolysin O (ASO) latex screening test with the ASO hemolytic test. Agreement between the two tests was poor when Difco streptolysin O (SLO) reagent was employed in the hemolytic test; approximately 34% of the sera with ASO titers in the normal range of the hemolytic test gave false-positive latex test reactions. However, the percentage of false-positive latex test reactions was only 5% when Behringwerke SLO reagent was used in the hemolytic test. An assay of the Difco and Behringwerke SLO reagents against an ASO standard indicated that the Difco SLO reagent was more potent than the Behringwerke SLO reagent. The lack of agreement between the Behringwerke latex test and the hemolytic test using Difco SLO reagent is attributed to the potency of the SLO reagents.     

Simple solutions to false-positive staphylococcal enterotoxin assays with seafood tested with an enzyme-linked immunosorbent assay kit (TECRA).

The TECRA kit, a commercial staphylococcal enterotoxin visual immunoassay kit, is an enzyme-linked immunosorbent assay system which utilizes polyvalent antisera against staphylococcal enterotoxin types A to E. The test is simple and rapid to perform (4 h) and has therefore been widely used for screening purposes. In this study, the TECRA kit produced a number of false-positive reactions with seafood; 25% of 218 samples of seven types of seafood gave false-positive results, particularly shellfish such as mussels (85%), clams (32%), oysters (23%), winkles (20%), and squid (13%). Some nonshellfish samples also gave false-positive results with the TECRA kit (smelt [20%] and trout [10%]). The substance contributing to the false-positive results differed from true staphylococcal enterotoxins in that it was: (i) heat labile, being completely inactivated by heating for 3 min at 70 degrees C, compared with 5% inactivation of true staphylococcal enterotoxins by the same heat treatment, (ii) in a selective reaction with normal rabbit or calf serum (nonspecific reactions were completely abolished by these sera, whereas staphylococcal enterotoxins were not affected), and (iii) incapable of binding to a copper-chelate Sepharose gel (all of the substance remained in the unbound wash fraction, whereas staphylococcal enterotoxins were quantitatively bound to the gel). The false-positive reactions occurring with seafood were not associated with substances produced by microorganisms, since the bacterial isolates from the samples did not give positive results with the TECRA kit.(ABSTRACT TRUNCATED AT 250 WORDS)     

Limitations of immunofluorescence tests in the diagnosis of infectious mononucleosis

The relative value of heterophil agglutinins (HA) and of specific EBV antibodies in the diagnosis of infectious mononucleosis (IM) was assessed in 108 cases of the disease and in 280 controls. Among the 108 cases 93 were HA-positive by sheep cells in at least one of their sera, while 15 were HA-negative by the same test. Among the 280 controls false-positive HA tests were not encountered except in eight cases with the horse cell microtitre tests. With one of the two slide tests at least two false-positive tests and 12 false-negative tests were also found but these sera had low titres in microtitre tests. The HA life-span was found to be unexpectedly long in a few cases, sheep cell HA lasting up to 8 to 10 months and horse cell HA up to 21 to 23 months.Many false-positive tests may therefore not be true false-positives and may result from the persistence of HA following unrecognized mononucleosis months before. Virtually all cases of IM had (or developed) antibodies to Epstein-Barr virus, viral capsid antigen (EBV-VCA), whereas only half of the controls were EBV-VCA-positive. The comparative analysis of nonspecific and specific test results in mononucleosis allows the following conclusions: (1) horse cell microtitre tests and the monospot test are more sensitive than sheep cell microtitre tests and the monotest; (2) false-negative results are occasionally seen with the latter tests but not with the former; (3) more false-positive results, however, are probably seen with the former tests; and (4) specific EBV-IgM and EBV-EA antibody tests are useful in the diagnosis of selected borderline cases of mononucleosis.     

Achieving High Accuracy Prediction of Minimotifs

The low complexity of minimotif patterns results in a high false-positive prediction rate, hampering protein function prediction. A multi-filter algorithm, trained and tested on a linear regression model, support vector machine model, and neural network model, using a large dataset of verified minimotifs, vastly improves minimotif prediction accuracy while generating few false positives. An optimal threshold for the best accuracy reaches an overall accuracy above 90%, while a stringent threshold for the best specificity generates less than 1% false positives or even no false positives and still produces more than 90% true positives for the linear regression and neural network models. The minimotif multi-filter with its excellent accuracy represents the state-of-the-art in minimotif prediction and is expected to be very useful to biologists investigating protein function and how missense mutations cause disease.     

Minimizing false positives in kinase virtual screens

In spite of recent improvements in docking and scoring methods, high false-positive rates remain a common issue in structure-based virtual screening. In this study, the distinctive features of false positives in kinase virtual screens were investigated. A series of retrospective virtual screens on kinase targets was performed on specifically designed test sets, each combining true ligands and experimentally confirmed inactive compounds. A systematic analysis of the docking poses generated for the top-ranking compounds highlighted key aspects differentiating true hits from false positives. The most recurring feature in the poses of false positives was the absence of certain key interactions known to be required for kinase binding. A systematic analysis of 444 crystal structures of ligand-bound kinases showed that at least two hydrogen bonds between the ligand and the backbone protein atoms in the kinase hinge region are present in 90% of the complexes, with very little variability across targets. Closer inspection showed that when the two hydrogen bonds are present, one of three preferred hinge-binding motifs is involved in 96.5% of the cases. Less than 10% of the false positives satisfied these two criteria in the minimized docking poses generated by our standard protocol. Ligand conformational artifacts were also shown to contribute to the occurrence of false positives in a number of cases. Application of this knowledge in the form of docking constraints and post-processing filters provided consistent improvements in virtual screening performance on all systems. The false-positive rates were significantly reduced and the enrichment factors increased by an average of twofold. On the basis of these results, a generalized two-step protocol for virtual screening on kinase targets is suggested.     

Increased risk of breast cancer in women with false-positive test: The role of misclassification

IntroductionStudies have shown that women with a false-positive result from mammography screening have an excess risk for breast cancer compared with women who only have negative results. We aimed to assess the excess risk of cancer after a false-positive result excluding cases of misclassification, i.e. women who were actually false-negatives instead of false-positives.MethodWe used data from the Copenhagen Mammography Screening Programme, Denmark. The study population was the 295 women, out of 4743 recalled women from a total of 58,003 participants, with a false-positive test during the screening period 1991–2005 and who later developed breast cancer. Cancers that developed in the same location as the finding that initially caused the recall was studied in-depth in order to establish whether there had been misclassification.ResultsSeventy-two cases were found to be misclassified. When the women with misclassified tests had been excluded, there was an excess risk of breast cancer of 27% (RR = 1.27, 95% confidence interval (CI), 1.11–1.46) among the women with a false-positive test compared to women with only negative tests. Women with a false-positive test determined at assessment had an excess risk of 27%, while false-positives determined at surgery had an excess risk of 30%.ConclusionsThe results indicate that the increased risk is not explained only by misclassification. The excess risk remains for false-positives determined at assessment as well as at surgery, which favours some biological susceptibility. Further research into the true excess risk of false positives is warranted.     

四维子宫输卵管超声造影假阳性与假阴性的原因分析

目的:调查与分析四维子宫输卵管超声造影假阳性与假阴性的原因。方法:选择2015年6月到2019年8月在本院妇产科临床初步诊断为输卵管不孕症患者125例,所有患者都给予X线子宫输卵管碘油造影(Hysterosalp ingography,HSG)与四维子宫输卵管超声造影,记录诊断效果、不良反应,判断假阳性与假阴性的发生原因。结果:在125例患者中,HSG诊断输卵管通畅33例,通而不畅72例,阻塞20例;四维超声造影诊断为输卵管通畅33例,通而不畅74例,阻塞18例。将HSG检查作为金标准,四维超声造影检查输卵管阻塞准确率93.6%,Kappa值=0.929(P<0.05),出现假阳性与假阴性共8例。四维超声造影检查期间发生的阴道少量出血、造影剂过敏、恶心呕吐、腹痛等不良反应发生率为2.4%,显著低于HSG检查的13.6%(P<0.05)。单因素分析结果显示合并糖尿病、产次、初潮年龄、孕次与四维子宫输卵管超声造影的假阳性与假阴性显著相关(P<0.05);多因素Logistic回归分析结果显示合并糖尿病、产次、初潮年龄为导致四维子宫输卵管超声造影假阳性与假阴性的主要原因(P<0.05)。结论:四维子宫输卵管超声造影可实时动态观察输卵管通畅情况,应用安全性也比较好,但是也存在假阳性与假阴性情况,合并糖尿病、产次、初潮年龄为导致四维子宫输卵管超声造影假阳性与假阴性的主要原因。     

Accurate decisions in an uncertain world: collective cognition increases true positives while decreasing false positives

In a wide range of contexts, including predator avoidance, medical decision-making and security screening, decision accuracy is fundamentally constrained by the trade-off between true and false positives. Increased true positives are possible only at the cost of increased false positives; conversely, decreased false positives are associated with decreased true positives. We use an integrated theoretical and experimental approach to show that a group of decision-makers can overcome this basic limitation. Using a mathematical model, we show that a simple quorum decision rule enables individuals in groups to simultaneously increase true positives and decrease false positives. The results from a predator-detection experiment that we performed with humans are in line with these predictions: (i) after observing the choices of the other group members, individuals both increase true positives and decrease false positives, (ii) this effect gets stronger as group size increases, (iii) individuals use a quorum threshold set between the average true- and false-positive rates of the other group members, and (iv) individuals adjust their quorum adaptively to the performance of the group. Our results have broad implications for our understanding of the ecology and evolution of group-living animals and lend themselves for applications in the human domain such as the design of improved screening methods in medical, forensic, security and business applications.     

Field test and experimental use of CYBEST model 2 for practical gynecologic mass screening

CYBEST is an automated cytologic screening system for uterine cancer utilizing a pattern-recognition image-analysis system. The prototype was developed in 1972 following fundamental studies of feature extraction, feature evaluation using ambiguity differential functions and segmentation of cell and nuclear images. Model 2 was developed in 1974 with an improved mechanism and function. The parameters employed are nuclear size, nuclear optical density, N/C ratio and nuclear shape. The data of field tests using 220 samples containing three cases of dysplasia, 110 cases of carcinoma and 107 nonmalignant cases were as follows: two false-negative cases (1.8%), 13 false positives (12.1%) and one reject (0.9%). This system was experimentally tested for practical mass population screening with 1,829 cases including 17 atypical cases (four epidermoid carcinomas). The data were as follows: no false-negative cases and 581 false-positive cases (32.1%). Of the latter, 311 cases (17.2%) were pathologic samples, such as severe cervicitis, senile colpitis, Trichomonas infestation, etc., and the remaining 270 cases (14.9%) were within physiologic limits, corresponding to true false-positive samples.     

Hepatitis B surface antigen: regional variation in sub-type ratio in the Canadian Red Cross donor population.

A total of 344 sera positive for hepatitis B surface antigen from volunteer blood donors at several Canadian Red Cross centres were subtyped for ad and ay specificity by counterelectrophoresis. Of the 50 sera from Toronto 21 (42%) were ad and 29 (58%) were ay; of the 95 from Montreal 82 (86%) were ad and 13 (14%) were ay; of the 199 from Quebec 179 (90%) were ad and only 20 (10%) were ay. The w and r specificities were also determined in 125 of the samples: 123 were w; the 2 samples of r specificity were from Toronto. On the other hand, among 45 sera from patients with acute hepatitis type B in Quebec 13 (29%) were ad and 33 (71%) ay.     

Echinococcus multilocularis: purification and characterization of glycoprotein antigens with serodiagnostic potential for canine infection

We show that a conventionally purified glycoprotein component of Echinococcus multilocularis protoscolex, designated as Emgp-89, may be useful as a serodiagnostic antigen for detecting E. multilocularis infection in dogs domesticated in endemic areas. Emgp-89 was obtained from the parasite material by a simple procedure using Con A-agarose and subsequent gel filtration chromatography. The purified fraction showed a molecular weight of >4000 kDa upon gel filtration and reacted with a series of lectins that specifically bind to mannose, galactose, N-acetylglucosamine, and N-acetylgalactosamine. Subsequently, serodiagnostic performance of Emgp-89 was evaluated through enzyme-linked immunosorbent assays (ELISAs) by using sera from normal, domestic dogs and dogs infected with other helminths. Emgp-89 positively reacted with all 16 serum samples from E. multilocularis-infected dogs, thus showing that this antigen is highly sensitive. On the other hand, the specificity of Emgp-89-based ELISA, determined using 41 serum samples from dogs infected with other helminths, was relatively low (83%). As an attempt to improve the specificity of Emgp-89-based ELISA, we pretreated Emgp-89 with proteinase K or sodium periodate, expecting that these treatments would enable discrimination of true positives from false positives. The ELISA value increased after treatment with sodium periodate in most false-positive samples, whereas significant decreases were observed in sera from all dogs infected with E. multilocularis. Further evaluation of this antigen should be performed using sera from dogs infected with closely-related parasites, including taeniid cestodes, which are expected to prove that this serodiagnostic system is sufficiently specific for clinical and field applications.     

Evaluation of Passive Hemagglutination, Solid-Phase Radioimmunoassay, and Immunoelectroosmophoresis for the Detection of Hepatitis B Antigen

Sensitivity and specificity of passive hemagglutination (RCA), solid phase radioimmunoassay (RIA), and immunoelectroosmophoresis (IEOP) were compared under experimental and clinical conditions. In dilution experiments with sera containing hepatitis B antigen (HB Ag) of known subtypes, the sensitivity for an ad subtype serum was RIA (1), RCA (1/2), IEOP (1/256) and for an ay subtype serum RCA (1), RIA (1/8), IEOP (1/128). An evaluation of the National Institutes of Health, Division of Biologics Standards test panel number 2 demonstrated HB Ag in 34 of 60 samples by RIA, in 33 by RCA, and in 25 by IEOP. HB Ag was detected in 57.5% of 200 outpatients with a tentative diagnosis of hepatitis by RIA, in 54% by RCA, and in 42.5% by IEOP. In 1,661 volunteer blood donors, 13 (0.78%) were "positive" for HB Ag by RIA, 11 (0.66%) by RCA, and 3 (0.18%) by IEOP. However, absorption experiments indicated that at least six of the above RIA positive and five of the RCA positive sera exhibited nonspecific positive reactions.     

Evaluation of five hepatitis C virus screening tests and two supplemental assays: performance when testing sera from sexually transmitted diseases clinic attendees in the USA

The performances of five screening tests (recombinant peptide-based first and second generation tests from Abbott and Ortho, and a synthetic peptide-based test from Biochem Immunosystems) and two supplemental tests: recombinant peptide- based, Abbott neutralization test and Chiron second generation recombinant immunoblot assay (RIBA 2), were evaluated for their ability to detect hepatitis C virus (HCV) antibodies in a population of 276 individuals attending a sexually transmitted diseases (STD) clinic in the USA. Although the five screening tests produced a variable number (35-62) of repeatedly reactive samples, only 13% (36/276) were classified as true positives by the supplemental tests. Thirty-four of the 36 were reactive by all screening tests and 32 of the true positives were reactive by both supplemental tests, while 2 did not neutralize but were reactive in the RIBA 2 test. Of the remaining 2 of the true positives which were discordant by several of the screening assays, 1 was confirmed by both supplemental assays but the other required a chemiluminescent enhancement technique to show positivity in RIBA 2. The sensitivities of the first and second generation Abbott and Ortho tests ranged from 97% to 100% and that of the Biochem test was 94%. The specificities of these tests ranged from 89.2% to 99.6%. The second generation Ortho test presented 9.4% (26/276) false positives. The use of second generation Ortho as a screening test would lead to an excessive number of confirmatory false positives. the positive predictive values of the screening tests ranged from 58.1% to 97.1%. Although the synthetic peptide based Biochem test exhibited the best overall indices, the presence of 2 false negative results would prevent its use as a singular screening test. Nevertheless its high specificity may lend itself to be used as a second screening test before confirmatory testing with RIBA 2.     

HIV-2 antibody testing of blood donors with doubtful immunoblot results for HIV-1

Antibodies against human immunodeficiency virus type-1 (HIV-1) in samples from blood donors are commonly detected by various enzyme-linked immunosorbent assays (ELISA) and by confirmatory tests, e.g., "Western blot" or immunofluorescence tests. Immunoblot reactivity, which is directed only towards the HIV-1 core proteins p 18, p 24 and p 55, may represent false-positive reactions. Out of 125,000 blood donations, 140 were repeatably HIV-1 antibody reactive by ELISA; of these, 20 were doubtful positive sera with isolated p 18 and/or p 24 bands in the HIV-1 confirmatory assay. Antibodies to HIV-2 are known to cross-react with these HIV-1 core proteins. We therefore assayed the 20 sera by immunofluorescence and immunoblotting for the presence of antibodies to HIV-2. None of these doubtful HIV-1 antibody positive blood donor sera was found to have antibodies to HIV-2.     

Evaluation of various radioimmunological techniques for the detection of antigen HBs]

The different radioimmunoassay technics routinely used for detection of HBs Antigen are Ausria I, Ausria II, RIA and CLB/RIA. Classification of these techniques either liquid or solide phase, is realized by nature of the tracer (Antigen or Antibody radioiodinated). The sensitivity and the specificity are presented as the comparison with other reactions. There is no significative difference on 1089 tested samples between Ausria I and RIA. The percentage of false positives in RIA or CLB/RIA reactions is 1.5% when the incubation is at 20 degrees C and 1% when it is at 37 degrees C. Advantages, disadvantages, cost prices, automation and possibility of computer treatment are discussed. The utilization of reference sera is proposed and the problem of the utilization en France of the radioactive products is approached.     

Preferred analysis methods for Affymetrix GeneChips revealed by a wholly defined control dataset

Background

As more methods are developed to analyze RNA-profiling data, assessing their performance using control datasets becomes increasingly important.

Results

We present a 'spike-in' experiment for Affymetrix GeneChips that provides a defined dataset of 3,860 RNA species, which we use to evaluate analysis options for identifying differentially expressed genes. The experimental design incorporates two novel features. First, to obtain accurate estimates of false-positive and false-negative rates, 100-200 RNAs are spiked in at each fold-change level of interest, ranging from 1.2 to 4-fold. Second, instead of using an uncharacterized background RNA sample, a set of 2,551 RNA species is used as the constant (1x) set, allowing us to know whether any given probe set is truly present or absent. Application of a large number of analysis methods to this dataset reveals clear variation in their ability to identify differentially expressed genes. False-negative and false-positive rates are minimized when the following options are chosen: subtracting nonspecific signal from the PM probe intensities; performing an intensity-dependent normalization at the probe set level; and incorporating a signal intensity-dependent standard deviation in the test statistic.

Conclusions

A best-route combination of analysis methods is presented that allows detection of approximately 70% of true positives before reaching a 10% false-discovery rate. We highlight areas in need of improvement, including better estimate of false-discovery rates and decreased false-negative rates.     

Immunodiagnosis of sexually transmitted disease

Methods for detecting microbial antigens in clinical specimens offer an alternative to culture in the diagnosis of some sexually transmitted diseases. Developers of the immunologic methods are faced with a number of problems in evaluating the new tests. Traditionally, these tests are compared to culture as the "gold standard." Unfortunately, culture for Neisseria gonorrhoeae or Chlamydia trachomatis--the two agents most commonly sought--is considerably less sensitive than 100 percent. Immunologic methods may appear to produce false positives when the paired specimens are actually false-negative cultures. Another source of discordant results is sampling variation. These considerations, however, will not account for all false-positive results. Even the best non-culture methods have a low rate of false-positive results. If a new test has a specificity of 97 percent, it, by definition, yields approximately 3 percent false-positive reactions. In low-prevalence settings this false-positive rate will create problems in interpreting the results. For example, in a population with 3 percent prevalence of infection, a positive result in a 97 percent specificity test could only have a predictive value of 50 percent. Most testing for STD agents is performed in low-prevalence settings. None of the currently available immunodiagnostic procedures has a performance profile that suggests it will be satisfactory for diagnostic use in the low-prevalence setting.     

Detection and Identification of Diagnostic Histoplasma capsulatum Precipitates by Counterelectrophoresis

Studies were carried out to develop and evaluate a counterelectrophoresis (CEP) technique for the rapid and specific identification of the diagnostically important histoplasmosis H and M precipitin bands. Well-defined and centrally located precipitin bands were produced by using a discontinuous buffer system and a gel matrix composed of agarose and ionagar no. 2. A template was devised which allowed the selective identification of the H and M precipitins. Comparative evaluations were performed with the microimmunodiffusion (ID) and complement fixation tests. In 52 sera from persons with histoplasmosis, either the H or M precipitin, or both, were identified in 42 (81%) of the cases with the CEP technique and in 43 (83%) with the ID test. With sera from 28 persons with heterologous diseases, the CEP technique, like the ID test, failed to react. The specificity of the CEP technique was dependent upon the use of the identity template. The CEP technique is recommended for routine use in laboratories testing moderate numbers of sera. It provides accurate and reproducible results within 90 min, in contrast to the ID test, which requires 18 to 24 h.     

Urease Color Test Medium U-9 for the Detection and Identification of “T” Mycoplasmas in Clinical Material

A urease color test fluid medium (U-9) for the detection and identification of T (T-strain) mycoplasmas in clinical material is described which is sensitive and specific for this group of mycoplasmas. The medium was prepared from commercially available components and contained 95% half-strength, tryptic digest broth (pH 5.5), 4% unheated horse serum, 0.05% highest-purity urea, 0.001% sodium phenolsulfonphthalein, and 1,000 units of potassium penicillin G per ml. The final reaction of medium U-9 was pH 6.0. The overall agreement (positive and negative) between urease reactions in U-9 urease color test medium and culture findings in a standard agar primary culture system among 686 clinical specimens was 98.1%. The disagreement consisted of 13 false-positive urease reactions which were recognized visually as false-positive reactions due to other microorganisms. For specimens from the female genitourinary tract, the inclusion of 2.5 mug of amphotericin B (Fungizone) per ml of medium U-9 is recommended for the suppression of growth of Candida species and filamentous fungi.