Maximum Likelihood Estimation of Titer via a Power Family of Four-Parameter Logistic Model.

For many laboratory assays, the readouts are presence or absence of a particular function, and the binary outcomes are correlated. The research interest is often focused on the estimation of titers, at which 50% positivity occurs. The classical approach by Reed and Muench (RM) assumes linear dose-response relationship around the potential titer, and uses only information from two points around the potential titer, which is inefficient in both precision and accuracy. While the model-based methods such as four-parameter logistic regression (4PL) use all the data, they do not consider the correlation among binary outcomes from same identities, which may lead to estimates with overstated precision. We propose estimating titers from two different anchors: independent responses from same identities or exchangeable responses from same identities. Marginal distributions of responses are linked to covariates of dilution factors by the 4PL model for independent responses and by a power family of the 4PL models for exchangeable responses. The maximum-likelihood procedure is used to get estimates of parameters and titers. The superiority of proposed methods over the classical approach is demonstrated both in simulation studies and in analysis of real data from hemagglutination assays.

Comparison of a Novel Test (ODAK Brucella Coombs Gel Test) with Commonly Used Serological Tests in Human Brucellosis.

BACKGROUND: Since Brucellosis is difficult to diagnose based on clinical symptoms, the diagnosis mostly relies on the results of serological testing. ODAK Brucella Coombs Gel Test is a novel and rapid gel microcolumn agglutination test which is performed in microcolumns containing gel matrix and Coombs antibodies. In this study, we aimed to compare ODAK Brucella Coombs Gel Test with other commonly used serological tests. METHODS: 150 blood samples of patients, preliminarily diagnosed as Brucellosis, were included in this study. Rose Bengal (RB), ODAK Brucella Coombs Gel Test (CGT), Brucellacapt (BCAP), and Standard Agglutination Test (SAT) were performed for all samples. Also, Coombs Agglutination Test (CAT) was performed for all SAT negative samples. 1/160 and above titers were accepted as positive result except RB which is a qualitative test. RESULTS: 100 (67%) out of 150 samples were found positive by RB. All of the 50 RB negative samples were also found negative by SAT and CAT test. However, 2 (4%) and 7 (14%) of them were positive by CGT and BCAP tests, respectively. Additionally, among 100 RB positive samples, only 68, 77, and 87 were positive by SAT+CAT combination, CGT, and BCAP tests, respectively. CONCLUSIONS: Currently, CGT is the only rapid (< 1 hour) serological test in which Coombs antibodies are used. Our results showed that negative results of RB, as a screening test, are not reliable enough as compared to CGT. However, positive RBT results confirmed with SAT were almost always, in most of the cases with higher titers, positive with CGT and BCAP. On the other hand, even if SAT is found negative with RB positivity, samples still must be investigated with CAT, CGT or BCAP. Consequently, CGT may be used as a rapid screening test instead of RB and it furthermore has similar sensitivity with the other confirmation tests in which Coombs antibodies are used. Therefore, ODAK Brucella Coombs Gel Test seems to be a very useful diagnostic tool for Brucellosis.

Is pretest probability assessment on emergency department patients with suspected venous thromboembolism documented before SimpliRED D-dimer testing?

BACKGROUND: The SimpliRED D-dimer assay is commonly ordered by emergency physicians for suspected pulmonary embolus or deep venous thrombosis. A pretest probability (PTP) assessment is required for the results of this diagnostic test to be interpreted correctly and applied appropriately. Without this assessment, the physician may misinterpret the test results and proceed to unnecessary diagnostic imaging (DI) or inappropriate discharge. Our objectives were to measure the documentation rate of PTP for emergency department (ED) patients on whom a SimpliRED D-dimer assay was performed for suspected venous thromboembolism (VTE) and to determine if the clinical management decisions that followed were in keeping with current recommendations. METHODS: In this medical record review, we used a random number generator to select 100 charts from all 760 patients who had a SimpliRED D-dimer performed during a 3-month period at an academic tertiary care centre with 3 EDs. Trained data abstractors, blinded to the study hypothesis, abstracted explicitly defined data from each chart. An independent abstractor assessed the reliability of 15 of the charts that were randomly chosen. RESULTS: Suspicion of VTE was documented in 97 of the 100 charts. There was no documentation of PTP assessment for 62 of the 97 cases. Ten had a positive D-dimer but 5 of these had no evidence of subsequent DI. Of the 97 charts reviewed, 24 documented decisions were in discordance with published clinical management recommendations for VTE. CONCLUSION: In the majority of ED cases of suspected VTE, PTP assessment was not documented and approximately one-quarter of these documented decisions were in discordance with established recommendations for the given test results. This suggests that PTP assessments are not being conducted in a significant proportion of cases and the diagnostic test results are misinterpreted, applied incorrectly or both.

Liver cystic echinococcosis: which cysts are correlated with false negative indirect passive hemagglutination (IHA)?

BACKGROUND: It's generally agreed that calcified liver cystic Echinococcosis (CE) is associated with negative serology; the influence of other cystic features on serologic result remains unclear. The aim of this study is to determine predictive factors of negative indirect passive hemagglutination (IHA) in liver CE. METHODS: 119 patients operated on for liver CE, had preoperative IHA. Correlation was studied between serological result and sex, age of patient and number (single vs multiple), size, ultrasonic type (unilocular, multivesicular, degenerative) and intrabiliary rupture of cyst. RESULTS: IHA sensitivity was 74.8%. Univaried analysis showed that false negative serology was correlated with age <10 or >20 years (p < 0.01), single cyst (p < 0.006), cystic size <9 cm (p < 0.03) and unilocular or degenerative cyst in comparison with multivesicular type (p < 0.01 and p < 0.03 respectively). IN CONCLUSION: in liver CE, cystic predictive factors of false negative IHA are single cyst, unilocular or degenerative type and size inferior to 9 cm.

Development of passive haemagglutination (PHA) and haemagglutination inhibition (HAI) technique for potency estimation of Cobra Antisnake Venom Serum (ASVS).

Antibodies against snake venom or antivenom potency are assayed quantitatively by in-vivo neutralization test in mice, which requires large number of laboratory animals. In potency assays of biological substances such as antivenoms, it is highly desirable to avoid suffering and death of animals by substituting in-vivo method with in-vitro methods, provided such methods measure life-saving capability with precision similar to that of in-vivo method. The in-vitro tests determine the neutralizing power of antivenom by permitting the evaluation of a particular biological activity of the venom and its neutralization after mixing the venom with the antivenom [Theakston RDG, Reid HA. Development of simple standard assay procedures for the characterization of snake venom. Bull WHO 1983;61:949-956; Gutierrez JM, Rojas G, Lomonte B, Gene JA, Chaves F, Alvarado J, et al. Standardizing of assays for testing the neutralizing ability of antivenoms. Toxicon 1990;28:1127-1129; Theakston R.D.G. Comments on letter of Gutierrez et al. on standardization of assays for testing the neutralizing ability of antivenoms. Toxicon 1990;28:1131-1132; Harvey AL, Barfaraz A, Thomson E, Faiz A, Preston S, Harris JB. Screening of snake venom for neurotoxic and myotoxic effects using simple in-vitro preparation from rodents and chicks. Toxicon 1994;32:257-265; World Health Organization Progress in characterization of venom and standardization of anti-venoms. Geneva: WHO offset publication; 1981. p. 58.]. Hence, the ideal requirements for an assay in detecting venom and venom antibody include high level of sensitivity, specificity (ability to differentiate between venom and venom antibody produced by closely related species of snakes), reproducibility and simplicity. A new in-vitro procedure for quantitative analysis of potency of ASVS by passive haemagglutination (PHA) and haemagglutination inhibition (HAI) has been explored. The methods described are simple, rapid, economical, reproducible and useful in replacing the more expensive in-vivo neutralization assays. Moreover, it also eliminates the use of laboratory animals.

Different accuracies of rapid enzyme-linked immunosorbent, turbidimetric, and agglutination D-dimer assays for thrombosis exclusion: impact on diagnostic work-ups of outpatients with suspected deep vein thrombosis and pulmonary embolism.

The requirement for a safe diagnostic strategy should be based on an overall posttest incidence of venous thromboembolism (VTE) of less than 1%, with a negative predictive value of more than 99 to 100% during 3-month follow-up. Compression ultrasonography (CUS) and spiral computed tomography (CT) currently are the methods of choice to confirm or rule out deep venous thrombosis (DVT) and pulmonary embolism (PE), respectively. CUS has a negative predictive value (NPV) of 97 to 98%, indicating the need to improve the diagnostic work-up of patients with suspected DVT by clinical score assessment and D-dimer testing. Spiral CT as a stand-alone method detects all clinically relevant PEs and a large number of alternative diagnoses. It rules out PE with a NPV of 98 to 99%. Spiral CT is expensive, emphasizing the need to improve the diagnostic work-up of patients with suspected PE by the use of clinical score assessment and D-dimer testing. Clinical score assessment for DVT and PE has not safely ruled out VTE in multicenter studies and in routine daily practices. Modification of the Wells clinical score assessment for DVT by elimination of the "minus 2 points" for alternative diagnosis will improve the reproducibility of the clinical score assessment. The combination of a first negative CUS and a negative SimpliRed or an enzyme-linked immunosorbent assay (ELISA) VIDAS D-dimer of < 1,000 ng/mL safely exclude DVT (NPV > 99%) irrespective of clinical score assessment and without the need to repeat CUS in approximately 60 to 70% of patients. The rapid quantitative and qualitative agglutination D-dimer assays for the exclusion of VTE are not sensitive enough as stand-alone tests and should be used in combination with clinical score assessment. A normal rapid ELISA VIDAS D-dimer test as a stand-alone test safely excludes DVT and PE, with a NPV of 99 to 100%, irrespective of clinical score, without the need of CUS or spiral CT. The combined strategy of a rapid ELISA VIDAS D-dimer followed by objective testing with CUS for DVT and by spiral CT for PE will reduce the need for noninvasive imaging techniques by 40 to 50%.

Further evaluation of a rapid diagnostic test for melioidosis in an area of endemicity.

Immunochromatographic test (ICT) kits for the rapid detection of immunoglobulin G (IgG) and IgM antibodies to Burkholderia pseudomallei were compared to the indirect hemagglutination (IHA) assay. In 138 culture-confirmed melioidosis cases, sensitivities were 80, 77, and 88% for IHA, ICT IgG, and ICT IgM, respectively. In a prospective study of 160 consecutive sera samples sent for melioidosis serology, respective specificities were 91, 90, and 69, positive predictive values were 41, 32, and 18, and negative predictive values were 99, 98, and 100%. ICT IgM kits are unreliable for diagnosis of melioidosis, but ICT IgG kits may be useful for diagnosing travelers presenting with possible melioidosis who return from regions where melioidosis is endemic.

Chagas' disease diagnosis: comparative analysis of recombinant ELISA with conventional ELISA and the haemagglutination test.

BACKGROUND AND OBJECTIVES: Serological screening for Chagas' disease in the blood banks of South America is carried out by using two different assays that generally show a high number of inconclusive results. To establish a combination of two tests that can minimize the number of inconclusive results, we compared a recombinant enzyme-linked immunosorbent assay (ELISA) with two conventional tests. MATERIALS AND METHODS: Serum samples from chagasic patients (n = 112), from non-chagasic individuals (n = 143) and from patients with other diseases (n = 32) were tested using three assays: recombinant ELISA (Rec-ELISA); conventional ELISA (Con-ELISA); and the indirect haemagglutination (IHA) test. RESULTS: When we evaluated the data by matching the Rec-ELISA and the IHA test, 52 inconclusive results were obtained. When Rec-ELISA and Con-ELISA were matched, only four inconclusive results were observed. CONCLUSIONS: Our investigation indicates that the use of two ELISAs with different antigen preparations provides an effective test combination for blood bank screening of Chagas' disease.

Prevalence of human parvovirus B19 in blood donors as determined by a haemagglutination assay and verified by the polymerase chain reaction.

BACKGROUND AND OBJECTIVES: Transmission of human parvovirus B19 (PV B19) by transfusion of blood and blood products is well documented. Although PV B19 infection is connected with severe complications in some recipients, donor screening is not yet mandatory. In this study the prevalence of PV B19, as detected by a haemagglutination assay (the Human PV B19 Antigen-Test), was assessed. In addition, the persistence of B19 DNA and the serological status of blood donors was also assessed. The specificity and utility of the Human PV B19 Antigen-Test for donor screening was investigated and compared with other screening strategies. MATERIALS AND METHODS: The prevalence of PV B19 viraemia was assessed in 28 972 donations from 15,660 remunerated donors by means of the haemagglutination assay. Reactive results were confirmed by the polymerase chain reaction (PCR). RESULTS: Overall, 255 donations gave reactive or indeterminate results in the screening assay. Four donations/donors detected by the haemagglutination assay were confirmed as positive for B19 DNA by PCR. Therefore, a frequency was detected of 1:7243 B19-positive donations and 1:3915 positive donors. Specificity was determined to be 99.1%. Follow-up showed the persistence of viraemia in low concentrations for prolonged time-periods. CONCLUSION: Blood donations with a high level of human PV B19 viraemia can be detected by the haemagglutination assay, which is rapid and easy to perform. The presence of neutralizing antibody may inhibit specific haemagglutination.

Invasive amoebiasis: an emerging parasitic disease in patients infected with HIV in an area endemic for amoebic infection.

OBJECTIVES: To describe the incidence and presentations of invasive amoebiasis (IA) in patients with HIV infection in an area endemic for amoebic infection and to assess the role of the indirect haemagglutination (IHA) assay in the diagnosis of IA in HIV-infected patients. DESIGN: Retrospective study of 18 cases of IA and HIV infection. SETTING: A university hospital, the largest centre for management of HIV-associated complications in Taiwan. METHODS: Medical, microbiological and histopathological records of 296 HIV-infected patients and serological data of IHA assay of 126 HIV-infected patients were reviewed to identify cases of IA from 23 June 1994 to 31 March 1999. An IHA titre > or = 1 : 128 was considered positive. Clinical characteristics of HIV-infected patients with IA and without IA were compared. RESULTS: Eighteen of the 296 patients (6.1%) with HIV infection were diagnosed with IA: 12 patients were diagnosed with definite IA and six with probable IA. The clinical manifestations included amoebic colitis (13 patients), amoebic liver abscess (nine), both colitis and abscess (four), and pleural effusion (two). IA was the initial presentation of HIV infection in nine patients. Co-infection with other enteric pathogens was diagnosed in six patients with IA. Compared with the 161 patients without IA who were newly diagnosed with HIV infection, the nine patients with IA had a higher median CD4+ lymphocyte count (202 x 10(6)/l versus 33 x 10(6)/l; P = 0.0017), were less likely to be diagnosed with AIDS (55.6% versus 85.4%; P = 0.039), and had fewer concurrent AIDS-defining illnesses (median number 0 versus 2; P = 0.003). Estimated mean survival duration was not significantly different between the two groups (597 days versus 611 days). Fourteen out of 126 patients (11.1%) had an IHA titre > or = 1 : 128. Of the 18 patients diagnosed with IA, 13 had a titre > or = 1 : 128. The sensitivity of IHA assay in the diagnosis of IA was 72.2% (13 out of 18) and the specificity was 99.1% (107 out of 108). The positive predictive value of IHA test for IA of this patient population was 92.9% (13 out of 14) whereas the negative predictive value was 95.5% (107 out of 112). CONCLUSION: IA is an increasingly important parasitic disease among patients with HIV infection in Taiwan. IHA assay has a good specificity and high negative predictive value in diagnosis of IA.

El diagnóstico serológico de la hidatidosis humana con ELISA y hemaglutinación indirecta / Serologic diagnosis of human hydatidosis

El diagnóstico serológico de la hidatidosis humana es fundamental ya que su detección temprana posibilita el éxito de la quimioterapia. El objetivo de este trabajo fue demostrar la validez del ELISA y la importancia de su asociación con la determinación del antígeno circulante. Se estudiaron 870 muestras por los métodos: HAI y ELISA para dosaje de anticuerpos (EAc) y antígeno circulante mediante un ELISA de captura (EAg). De los 870 sujetos estudiados 144 tenían hidatidosis. De los 144 sujetos con hidatidosis, veintiséis correspondían a sujetos operados en el período 1990-1993 observándose una buena correspondencia entre los resultados obtenidos por la HAI y el EAc. El resto (118) corresponden a sujetos con diagnóstico prequirúrgico por imágenes y sus resultados muestran que el EAc permitió detectar mayor número de portadores de quistes pulmonares (veintitrés) que la HAI (catorce), significando una mayor sensibilidad diagnóstica (Chi cuadrado: 9,8). La sensibilidad de estos métodos fue de 83 por ciento para la HAI y 91 por ciento para el EAc. Fueron detectados cuatro falsos negativos por EAc mediante el EAg lográndose as161 una sensibilidad del 95 por ciento al asociar ambos métodos. También mediante el EAg se logró detectar recidivas y no se observó influencia del tamaño de los quistes en la reactividad (AU)

El diagnóstico serológico de la hidatidosis humana con ELISA y hemaglutinación indirecta / Serologic diagnosis of human hydatidosis

El diagnóstico serológico de la hidatidosis humana es fundamental ya que su detección temprana posibilita el éxito de la quimioterapia. El objetivo de este trabajo fue demostrar la validez del ELISA y la importancia de su asociación con la determinación del antígeno circulante. Se estudiaron 870 muestras por los métodos: HAI y ELISA para dosaje de anticuerpos (EAc) y antígeno circulante mediante un ELISA de captura (EAg). De los 870 sujetos estudiados 144 tenían hidatidosis. De los 144 sujetos con hidatidosis, veintiséis correspondían a sujetos operados en el período 1990-1993 observándose una buena correspondencia entre los resultados obtenidos por la HAI y el EAc. El resto (118) corresponden a sujetos con diagnóstico prequirúrgico por imágenes y sus resultados muestran que el EAc permitió detectar mayor número de portadores de quistes pulmonares (veintitrés) que la HAI (catorce), significando una mayor sensibilidad diagnóstica (Chi cuadrado: 9,8). La sensibilidad de estos métodos fue de 83 por ciento para la HAI y 91 por ciento para el EAc. Fueron detectados cuatro falsos negativos por EAc mediante el EAg lográndose as161 una sensibilidad del 95 por ciento al asociar ambos métodos. También mediante el EAg se logró detectar recidivas y no se observó influencia del tamaño de los quistes en la reactividad

Clinical significance of measurements of antithyroid antibodies in the diagnosis of Hashimoto's thyroiditis: comparison with histological findings.

Measurements of antithyroglobulin and antimicrosomal (antiperoxidase) antibodies have been performed widely for the clinical diagnosis of autoimmune thyroid diseases. The present study was designed to compare these antibody titers with histological findings of the thyroid in patients with diffuse goiter who were suspected of having Hashimoto's thyroiditis. One hundred and ten euthyroid or hypothyroid patients (10 males and 100 females; age 48 +/- 15 (SD) years old) with diffuse goiter were studied for the measurement of antithyroglobulin and antimicrosomal or antiperoxidase antibodies by a hemagglutination technique (TGHA and MCHA, respectively) and by a newly developed radioassay (TgAb and TPOAb, respectively). The antibody titers were compared with the histological findings obtained by needle biopsy. TgAb, TPOAb, TGHA, and MCHA were detected in 80 (96.4%), 61 (73.5%), 37 (44.6%), and 54 (65.1%) of 83 patients with histologically proven Hashimoto's thyroiditis, respectively, but in only one (3.7%) of 27 patients without any inflammatory changes in the biopsy specimen. In 55 patients with negative TGHA and MCHA, the TgAb positivity was more closely associated with the histological diagnosis of Hashimoto's thyroiditis than the TPOAb positivity was, the incidence of each antibody in Hashimoto's thyroiditis being 89.7% (26/29) and 27.6% (8/29), respectively. In conclusion, the histological diagnosis of Hashimoto's thyroiditis can most precisely be predicted by the newly developed radioassay for TgAb.

[Comparison of rheumatoid test procedures--value and critical interpretation of sensitivity and specificity and their effect on pre-test and post-test probability]. / Rheumafaktortestverfahren im Vergleich--Aussagekraft und kritische Interpretation von Sensitivität und Spezifität sowie deren Einfluss auf Pre-Test- und Post-Test-Wahrscheinlichkeiten.

In this prospective study, sera of 440 patients with rheumatic and degenerative joint diseases were tested for the presence of rheumatoid factor (RF). The Latex agglutination test (LFT), Waaler-Rose hemagglutination, laser nephelometry and IgM-Enzyme immunoassay (IgM-EIA) were used for detecting IgM-rheumatoid factors. In addition, rheumatoid factor of IgA isotype was measured by an IgA-Enzyme immunoassay. Sensitivity, specificity, pre-test- and post-test-probability were evaluated based on the data obtained to compare the test systems used. Under prospective patient selection, none of the test systems used reached a sensitivity of 100% concerning its cut off level. Despite this limitation, latex agglutination and IgM-EIA reached the highest sensitivity. Waaler-Rose test (90,8%) showed the best result for specificity. The IgA-EIA held the third position in sensitivity, specificity and efficiency. By comparing sensitivity with specificity, no test system can be recognized as the absolutely best one, since the receiver operating characteristic curves (ROC) overlapped. Practically rheumatoid factor measurement should initially use a highly sensitive assay, such as LFT and IgM-EIA to screen for RF. In the case of a positive result a more specific assay should be used, for example laser nephelometry, to confirm the result.

Evaluación de una técnica ELISA para el despistaje de enfermedad de chagas: datos de seroprevalencia en la ciudad de Rosario / Evaluation of an ELISA for chagas disease screening: seroprevalence data in the city of Rosario

Se evaluó la performance de un ELISA para la detección de anticuerpos anti Trypanosoma cruzi, en comparación con las técnicas habitualmente usadas en este laboratorio (aglutinación de látex, aglutinación directa de parásitos coloreados, hemaglutinación indirecta e inmunofluorescencia indirecta). La comparación se realizó en forma simultánea y a doble ciego sobre 690 dadores voluntarios de sangre y 244 pacientes que asistían a consulta médica entre los meses de diciembre de 1992 y enero-febrero de 1993. La técnica en estudio resultó confiable tanto para el despistaje de dadores de sangre como para el diagnóstico de laboratorio. La seroprevalencia de la infección fue 8,1 por ciento entre los dadores de sangre y 12,3 por ciento entre los pacientes (AU)