Bicentric Evaluation of Six Anti-Toxoplasma Immunoglobin G automated. immunoassays and comparison with the LDBio-Toxo II Immunoglobulin G

Transcription

1 CVI Accepts, published online ahead of print on July 00 Clin. Vaccine Immunol. doi:./cvi Copyright 00, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved Full-Length Paper Bicentric Evaluation of Six Anti-Toxoplasma Immunoglobin G automated immunoassays and comparison with the LDBio-Toxo II Immunoglobulin G Western Blot. Arnaud Maudry, 1 Gautier Chene, Rémi Chatelain, 1 Hugues Patural, Bahrie Bellete, 1 Bernard Tisseur, Jamal Hafid, 1- Hélène Raberin, 1 Sophie Beretta, Roger Tran Manh Sung, 1- Georges Belot, and Pierre Flori 1-* 1 Pôle de Biologie-Pathologie, Parasitology and Mycology Laboratory, University Hospital of Saint Etienne-France Pôle Mère et Enfant,, University Hospital of Saint Etienne-France Laboratoire d analyses médicales Synerbio, Saint Etienne, France Unité d immunologie et de Physiologie, Faculté des Sciences et Techniques, Marrakech, Morocco Groupe immunité des muqueuses et agents pathogènes, GIMAP EA-0, Saint Etienne, France. *Corresponding author: Dr P. FLORI, 1 Pôle de Biologie-Pathologie, Laboratoire de Parasitologie et Mycologie, Hôpital Nord, CHU de Saint Etienne, 0 Saint Etienne, France. Phone: () 0 Fax: () pierre.flori@univ-st-etienne.fr Running titer: Evaluation of Anti-Toxoplasma IgG immunoassays. Key words : toxoplasmosis, Toxoplasma gondii, Immunoglobulin G, serological follow-up, immunoassays. Downloaded from on August 1, 01 by guest 1

2 Abstract A comparative study of the Toxoplasma IgG I and IgG II Access (Access I and II) (Beckman Coulter Inc, CA), Axsym Toxo IgG (AxSYM) (Abbot Diagnostics, IL), Vidas Toxo IgG (Vidas) (biomerieux, France), Immulite Toxo IgG (Immulite) (Siemens Healthcare Diagnostics Inc, IL) and Modular Toxo IgG (Modular) (Roche Diagnostics, Switzerland) tests was done on 0 consecutive serum samples. The Toxo II IgG Western Blot (LDBio, France) was done as a reference technique in case of inter-technique discordance. /0 sera were discordant among the different techniques. Using the 1 positive sera, we evaluated the standardization of the titrations obtained in IU/ml; The median ( nd quartile) obtained was.1 IU/ml for AxSYM, 1 IU/ml for Access I,. IU/ml for Access II, IU/ml for Vidas,. IU/ml for Immulite and IU/ml for Modular. For all the immunoassays tested, the following relative sensitivity and specificity values were found:.-0% for Access II,.-.% for Immulite, 0.-.% for AxSYM, 1.-.% for Vidas,.-.% for Access I and.-.% for Modular. Among the 0 sera, we did not find any false positive values with different tests for the same serum. Except for the Modular test which prioritized sensitivity, it appears that the positive cut-off values suggested by the pharmaceutical companies are very high (either for economical or for safety reasons). This led to an imperfect sensitivity, a large number of unnecessary serological follow-up of pregnant women and difficulty in determining the serological status of immuno-suppressed individuals. Downloaded from on August 1, 01 by guest

3 Introduction Toxoplasmosis, caused by Toxoplasma gondii, is widespread in human and warm-blooded animals. Although usually asymptomatic in immunocompetent humans, toxoplasmosis may cause severe disorders in pregnant women because of the high risk of transplacental transmission and the occurrence of abortion or multiple congenital lesions in the fetus and in immunocompromised individuals (, ). Life-threatening reactivation of a previous infection is commonly observed in cases of severe immunodeficiency (HIV infected patients, organ and haematopoietic stem cell transplant patients). For these patients, detection of Toxoplasma specific antibodies showing serological reactivation or primary infection are therefore essential for appropriate diagnosis and prevention of severe toxoplasmosis (, ). The follow-up of obstetrical toxoplasmosis mainly depends on the detection of antitoxoplasma specific Immunoglobulin M (IgM) and Immunoglobulin G (IgG) antibodies (1, 1, 1). The presence of toxoplasma specific IgM at the time of the first blood test is a cause for concern. The presence of toxoplasma specific IgG without IgM permits to confirm immunization of the patients and thus avoid unnecessary and expensive follow-up. In both, (Obstetrical follow-up and diagnosis in immunocompromised patients), IgG tests are crucial. Since the s, Toxoplasma IgG assays have been standardized by different generations of World Health Organization (WHO) standards (1) and test results have been reported in international units per milliliter (IU/ml). The Dye test (DT) first described by Sabin and Feldman 0 years ago, is still the reference method for the serodiagnosis of toxoplasmosis. However, this test uses live Toxoplasma gondii and is now only used in a few laboratories (1). A good alternative (the test Toxo II IgG Western Blot, LDBio, Lyon, France) has been proposed by Franck et al., () as a confirmatory technique: The results of this test appear to be consistent with those of the DT with a specificity of 0% and a sensitivity of.%. Thus, Downloaded from on August 1, 01 by guest

4 this immunobloting technique can be used as a very reliable and easy confirmatory test in laboratories where the DT cannot be implemented. In spite of International standardization and the availability of a reference (or confirmatory test), automated immunoassays frequently show discordance and moderate degree of correlation (, 1). A comparison of six random-access immunoassays (that report IgG levels in IU/ml) and the Toxo II IgG Western Blot (LDBio, Lyon, France) as a confirmatory technique was undertaken to review analytical performance and the degree of standardization. Materials and methods Samples This study was done on 0 consecutive sera from adult patients tested in our hospital laboratory (CHU de Saint Etienne, France) for 0 days in December 00. (.%) of these sera were from pregnant women who were followed up, (. %) were from patients followed up for immunodepression, and 1 (. %) sera came from other departments. As most sera were from pregnant women the M/F sex ratio was 1/.0. All the samples were given anonymous labels (as suggested by the ethical committee), decanted and frozen within hours of receipt. Then they were thawed and recentrifuged in series and tested in parallel in the first laboratory (Parasitology and Mycology Laboratory, University Hospital of Saint Etienne- France) on the same day with the Access Toxo IgG I, Access Toxo IgG II (Access I or Access II) (Beckman Coulter Inc, CA), Axsym Toxo IgG (AxSYM) (Abbot Diagnostics, IL) and Vidas Toxo IgG (Vidas) (biomerieux, Marcy l Etoile, France). The samples were frozen again immediately following testing and another freezing and thawing was done for testing in the second laboratory (Laboratory Downloaded from on August 1, 01 by guest

5 Synerbio,, rue Traversière, Saint Etienne, France) with the Immulite Toxo IgG (Immulite) (Siemens Healthcare Diagnostics Inc, IL) and Modular Toxo IgG (Modular) (Roche Diagnostics, Basel, Switzerland). In case of qualitative discordance between techniques (negative, equivocal or positive) the Western Blot was done as a confirmatory test Tests (i) Automated Immunoassays Six of the assays were automated using random-access instruments that could perform a range of infectious disease and biochemical assays. All tests were performed as instructed by the manufacturer. The manufacturer s cut off was applied to determine reactivity of the sample. All assays report the test results in IU/ml. The random-access immunoassays were listed previously. In Europe, Access II replaced Access I during the year 00. Access I (, ), AxSYM (,,, 1) and Vidas (,,, 1) use the WHO second international standard (IS) in contrast to Access II, Immulite () and Modular () which use the third WHO IS. It is to be noted that neither the second nor the third WHO IS has been tested in parallel with the six automated techniques in the present study. The main features of each automated test are described in Table 1. (ii) Western Blot () The Western Blot test is a qualitative immunoenzymatic test in which immunoblotting to nitrocellulose strips are used. After standardized incubation with sera and fixation of specific IgG on the band, antitoxoplasma IgG bound to the strip is detected using alkaline phosphatase-conjugated antibody and a specific substrate (nitroblue tetrazolium -bromo--chloro--indolylphosphate). The resulting bands on Downloaded from on August 1, 01 by guest

6 the patient s strip, corresponding to 0, 1,, 0 and kda. A positive result is defined by the presence of at least three matching bands on the patient s strip, including the specific band at 0 kda. A doubtful result is defined by the presence of the specific band at 0 kda with less than three matching bands on the patient s strip Analysis (i) Relative sensitivity, specificity, positive and negative predictive values (PPV and NPV) The relative sensitivities and specificities of each automated immunoassay were estimated by comparing the immunoassay s qualitative result (negative, equivocal and positive) with the result of the western blot. The relative sensitivity and the specificity of each immunoassay were calculated twice, first interpreting equivocal as negative and then as positive (Table ). PPVs and NPVs were calculated using the seroprevalence of our sampling (.1%) which is very close to the French seroprevalence (.%) (1). (ii) Statistical analyses The results expressed as IU/ml were used in the statistical analyses. Where an assay produced a result expressed as greater than the highest limit of the assay, the result was assigned a value of the limit level plus one. When, exact titer was not always determined, the range of the test s value was shown by the first, second and third quartile. Statistical assessment of differences of sensitivity, specificity, PPV and NPV with the different tests was estimated by Chi Square test (p <.0 was considered significant). Statistical assessment of differences of the mean titers with the different tests was estimated by ANOVA (Fisher-Snedecor test, p <.0 was considered significant). Downloaded from on August 1, 01 by guest

7 Results Global characteristics and concordance of the six automated immunoassays Out of 0 sera tested, 1 (.%) were negative with all the six tests, 1 (.%) were positive with all the six tests and (.%) were discordant. As a result the overall concordance of all six techniques was 1.% (1/0 sera). All the results of the discordant sera are shown in Table. Twenty two of these sera were positive by the Western Blot with, and specific bands corresponding to doubtful or false negative results with the majority of the automated tests. Of the remaining 1 sera, three had equivocal result with the Western Blot (with specific bands one of which was p0) and were negative with the Western Blot. The latter cases had false positive results (as compared to the Western Blot which was negative) but only with the AxSYM test (1 serum) and the modular test ( sera). Standardization of titrations (IU/ml) obtained with the six automated immunoassays Using the 1 positive sera which were confirmed by Western Blot, we evaluated the standardization of the titrations obtained (in IU/ml) by comparing their distribution (1 st, nd and rd quartiles); The median ( nd quartile) obtained from our sampling was statistically different from one test to the other, and varying from.1 IU/ml for AxSYM, 1 IU/ml for Access I,. IU/ml for Access II, IU/ml for Vidas,. IU/ml for Immulite and IU/ml for modular (Figure 1). In the same manner, (despite the bias caused by the different superior linearity limits of each technique), the means (+/- SE) obtained were. (±.),.1 (±.),.0 (±.0),. (±.1),. (±.),. (± 1.1) for the AxSYM, Access II, Access I, Vidas, Immulite and Modular respectively. Downloaded from on August 1, 01 by guest 1 Relative sensitivity, specificity, positive and negative predictive values (PPV and NPV)

8 The reliability of each test was determined and the characteristics are shown in Table. The only technique which showed a specificity of 0% was Access II. All the other tests had a specificity of.% except for the Modular test which showed a specificity less than.%. Regarding their sensitivities, when evaluating with the highest cut-off, it was found to be low with.,., 0., 1.,., and.% for Immulite, Access II, Axsym, Vidas, Access I and Modular test respectively. There was significant improvement in their sensitivities (p< 0.0, χ test) when the lowest cut-off for each technique was used: the sensitivities became.,.,,.,. and.% respectively in the same order; With the exception of the modular test, the latter sensitivities (with lowest cut-off) were associated with a specificity always greater than %. Discussion There were two objectives for this study: - To evaluate the standardization between the main automated tests. - To define the main reliability criteria (specificity, sensitivity, PPV, NPV) for each of the automated tests. Regarding the level of inter-technique standardization, the results were not as good as expected. This has been demonstrated in this study where the medians of the results varied by a factor of 1 to depending on the test used (Fig 1). These inter-technique variations are particularly surprising since in fact an international reference exists (1). In comparison, a recent study () evaluating anti-rubella virus IgG tests, of which six were automated, showed a much better standardization with a variation factor ranging between 1 to (average of the positive samples with different tests). We can speculate that these variations could be related to the differences in the antigenic solutions used by each of these tests. Indeed, Downloaded from on August 1, 01 by guest

9 the study by Petithory et al., 1 (1) showed variations related to the antigenic nature (complex antigenic structure with numerous membrane and cytosolic antigens) but at a much lower level. These different antigenic solutions induce an intense and precocious response in some cases and more delayed responses in other cases (, ). But the major cause of inter-technique variation is probably associated with the lack of cooperation of the companies providing the tests, thus limiting the harmonization and standardization of the titrations obtained and also the cut-offs proposed. The choice of the second or third WHO IS for standardisation of the different techniques could also contribute to the variability of the quantitative results. However, in our comparative study there was no signiificant difference between those which used the second and those which used the third WHO IS (Fig 1). On the other hand, it is interesting to note that the standards of the third WHO IS contain IgG with low avidity (1) and can as a result, influence the reactivity of the different techniques. In any case, all these factors induce considerable inter-technique variability and these variations especially for the AxSYM test and the Modular test as compared to the other tests (Fig1), make any comparison of inter-technique titrations impossible: To date, one can compare two successive titrations only if the latter are done using the same technique and preferably in the same series. It is interesting to note that despite the quantitative serologic variations, the number of qualitative discordant cases (negative, equivocal, positive) among the six automated techniques remained low. Thus, only /0 (.%) of the samples showed discordance (negative with one test and doubtful or positive with another) and only 1/ (.1%) with total discordance i.e. negative with one test and positive with another. This high concordance level is related to the variation in parallel of the cut-offs and the medians of the positive samples obtained from each of these techniques. For example, for the AxSYM IgG test, the cut-off was IU/ml and the median was.1 IU/ml and for the modular IgG test, the cut-off was 0 IU/ml and the median was IU/ml. These results which showed good concordance are in agreement with previous studies (,, 1) and confirm the reliability of all the automated techniques. Downloaded from on August 1, 01 by guest

10 In order to determine the criteria of the reliability of the different techniques, we have compared the sensitivities, specificities, PPV and NPV of the six tests. When the doubtful (or equivocal) values were considered as negative, the specificities were excellent: 0% for Access II,.% for Access I, AxSYM, Vidas and Immulite, and.% for modular (p> 0.0, no significant difference, χ test). It has to be said that, for the Modular test, the three false positive cases had an equivocal result by Western Blot not allowing to confirm the presence of specific antibodies. Therefore, these sera were not necessarily false positive. On the other hand, the sensitivities were very close and low for all the presently commercialysed tests and varied between.% to 1.%, with the exception of the Modular test which had a much higher sensitivity of.%. When the doubtful (or equivocal) values were considered as positive, the specificities remained excellent (> %) for all techniques except for the Modular technique and their sensitivities increased significantly : they were.%,.%, %,.%,.% for the tests Immulite, Access II, Vidas, Access I and AxSYM respectively (Table ). When the reliability criteria were analysed and also after analysing the receiver operating characteristics (ROC) (data not shown), we observe that almost all companies have prioritized specificity by raising the cut-offs. On the contrary, the Roche Company (Modular) has given priority to sensitivity by choosing a lower cut-off. Paradoxically, the latter technique has the most elevated cut-off with 0 IU/ml! These surprising results however are coherent with those of Franck et al. () which showed sensitivity and specificity levels which were totally different between the Vidas technique and the Cobas test (Cobas and Modular belong to the Cobas family tests ). For all the automated techniques studied, the sensitivities and specificities obtained should be analysed and interpreted in parallel with the quantitative results for the discordant and borderline sera (Table ). These results confirm the idea that the cut-offs chosen by the companies (for all the tests except for Modular) are too high and guarded. The lower cut-offs would have a better balance between sensitivity and specificity (Table ). We have also noticed that it is possible to have false positive cases even though Downloaded from on August 1, 01 by guest

11 1 at very low frequencies. An interesting point of our study is that there are not so many false positive or doubtful sera with these automated techniques. As a result, in case of a primary doubtful result with one automated technique, it seems advisable to test the serum using another automated technique since cross false positivity between techniques has not been found. Thus, in the present study, all the positive or doubtful sera with two automated techniques were confirmed to be either positive (0/ cases) or indeterminate (/ cases) by the Western Blot. Evidently, the possibility of doing a confirmatory test either by the Western Blot or the Dye Test would be more securing and would permit to avoid unnecessary and expensive serological follow up in up to % of cases. Acknowledgements We acknowledge the staff of the Parasitology-Mycology and Bacteriology-Virology laboratory of the Saint Etienne University Hospital France, for their skillful technical assistance in this study. Downloaded from on August 1, 01 by guest

12 References 1. Berger, F., V. Goulet, Y. Le Strat, and J. C. Desenclos. 00. Toxoplasmose chez les femmes enceintes en France : évolution de la séroprévalence et de l incidence et facteurs associés, Bull. Epidemiol. Hebd. 1:-. (in French). Derouin, F., H. Pelloux, on behalf of the ESCMID Study Group on Clinical Parasitology. 00. Prevention of toxoplasmosis in transplant Patients. Clin. Microbiol. Infect. 1:-.. Dimech, W., L. Panagiotopoulos, B. Francis, N. Laven, J. Marler, D. Dickeson, T. Panayotou, K. Wilson, R. Wootten, and E. M. Dax. 00. Evaluation of eight anti-rubella virus immunoglobulin G immunoassays that report results in international units per milliliter. J. Clin. Microbiol. :1-.. Flori, P., B. Bellete, C. Crampe, A. Maudry, H. Patural, C. Chauleur, J. Hafid, H. Raberin, and R. Tran Manh Sung. 00. A technique for dating toxoplasmosis in pregnancy and comparison with the Vidas anti-toxoplasma IgG avidity test. Clin. Microbiol. Infect. 1:-.. Foulon, W., I. Villena, B. Stray-Pedersen, A. Decoster, M. Lappalainen, J. M. Pinon, P. A. Jenum, K. Hedman, and A. Naessens. 1. Treatment of toxoplasmosis during pregnancy: A multicenter study of impact on fetal transmission and children s sequelae at age 1 year. Am. J. Obstet. Gynecol. :-1.. Franck, J., Y. J. Garin, and H. Dumon. 00. LDBio-Toxo II immunoglobulin G Western blot confirmatory test for anti-toxoplasma antibody detection. J. Clin. Microbiol. :-.. Fricker-Hidalgo, H., C. E. Bulabois, M. P. Brenier-Pinchart, R. Hamidfar, F. Garban, J. P. Brion, J. F. Timsit, J. Y. Cahn, and H. Pelloux. 00. Diagnosis of toxoplasmosis after allogeneic stem cell transplantation: Results of DNA detection and serological techniques. Clin. Infect. Dis. :e-e1. Downloaded from on August 1, 01 by guest 1

13 Goubet, S., H. Pelloux, H. Fricker-Hidalgo, A. Goullier-Fleuret, and P. Ambroise-Thomas. 1. Sérodiagnostic de la toxoplasmose : comparaison de la trousse Elisa AxSYM (Abbott) avec la trousse Vidas (BioMérieux), l'immunofluorence indirecte et l'isaga. Ann. Biol. Clin. :1-. (in French). Montoya, J. G., and O. Liesenfeld. 00. Toxoplasmosis. Lancet. :1-1.. Owen, W. E., T. B. Martins, C. M. Litwin, and W. L. Roberts. 00. Performance characteristics of six IMMULITE 000 TORCH assays. Am. J. Clin. Pathol. 1: Petersen, E., M. V. Borobio, E. Guy, O. Liesenfield, V. Meroni, A. Naessens, E. Spranzi, and P. Thulliez. 00. European multicenter study of the LIAISON automated diagnostic system for determination of Toxoplasma gondii-specific immunoglobulin G (IgG) and IgM and the IgG avidity index. J. Clin. Microbiol. : Petithory, J. C., P. Ambroise-Thomas, J. De Loye, H. Pelloux, A. Goullier-Fleuret, M. Milgram, C. Buffard, and J. P. Garin. 1. Serodiagnosis of toxoplasmosis: a comparative multicenter study of a standard scale through various actual tests and expression of the results in international units. Bull. W. H. O. :1-. (in French). 1. Reiter-Owona, I., E. Petersen, D. Joynson, H. Aspöck, M. L. Dardé, R. Disko, O. Dreazen, H. Dumon, R. Grillo, U. Gross, M. Hayde, R. Holliman, D. O. Ho-Yen, K. Janitschke, P. A. Jenum, K. Naser, M.Olszewski, P. Thulliez, and H. M. Seitz. 1. The past and present role of the Sabin-Feldman dye test in the serodiagnosis of toxoplasmosis. Bull. W. H. O. :-. 1. Remington, J. S., P. Thulliez, and J. G. Montoya. 00. Recent developments for diagnosis of toxoplasmosis. J. Clin. Microbiol. : Rigsby, P., S. Rijpkema, E. C. Guy, J. Francis, and R. G. Das. 00. Evaluation of a candidate international standard preparation for human anti-toxoplasma immunoglobulin G. J. Clin. Microbiol. :1-1. Downloaded from on August 1, 01 by guest 1

14 Roberts A, K. Hedman, V. Luyasu, J. Zufferey, M.H. Bessières, R.M. Blatz, E. Candolfi, A. Decoster, G. Enders, U. Gross, E. Guy, M. Hayde, D. Ho-Yen, J. Johnson, B. Lécolier, A. Naessens, H. Pelloux, P. Thulliez, and E. Petersen Multicenter evaluation of strategies for serodiagnosis of primary infection with Toxoplasma gondii. Eur. J. Clin. Microbiol. Infect. Dis. 0:-. 1. Roux-Buisson, N., H. Fricker-Hidalgo, A. Foussadier, D. Rolland, A. S. Suchel-Jambon, M. P. Brenier-Pinchart, and H. Pelloux. 00. Comparative analysis of the VIDAS Toxo IgG IV assay in the detection of antibodies to Toxoplasma gondii. Diagn. Microbiol. Infect. Dis. : Sensini A. 00. Toxoplasma gondii infection in pregnancy: opportunities and pitfalls of serological diagnosis. Clin. Microbiol. Infect. 1: Sickinger, E., F. Gay-Andrieu, G. Jonas, J. Schultess, M. Stieler, D. Smith, M. Hausmann, R. Stricker, R. Stricker, J. Dhein, and H. B. Braun. 00. Performance characteristics of the new ARCHITECT Toxo IgG and Toxo IgG Avidity assays. Diagn. Microbiol. Infect. Dis. :-. Downloaded from on August 1, 01 by guest 1 1

15 International Unit (logarithm scare) 00,0 0,0,0 1,0 00 IgG Axsym ,,0 1,0 1,1 1,0 IgG I Access 1 IgG II Access IgG Vidas Automated techniques IgG Immulite 0 IgG Modular 1 Figure 1. Distribution of the 1/0 Toxoplasma IgG positive sera confirmed by the LD Bio Toxo II IgG Western Blot technique. From bottom to top: The lowest value confirmed by at least bands (including p0) by the Western Blot technique; the first value in bold corresponds to the first quartile, 0,0 Downloaded from on August 1, 01 by guest the second value in bold to the median or second quartile, the third value in bold to the third quartile and the highest value corresponds to the limit of linearity of the technique without dilution. 1

16 Table 1. Characteristics of the Anti-Toxoplasma Immunoglobulin G automated immunoassays. Immunoassay Used on the following Robots Lower Cut off (IU/ml) Higher cut off (IU/ml) International Standard used for calibration Calibration curve Access Toxo IgG I Access Toxo IgG II Vidas Toxo IgG Not used since 00 Access II DXI 00 DXI 00 Mini Vidas Vidas WHO nd Multipoint.. WHO rd Multipoint WHO nd Two point AxSYM Toxo IgG AxSYM WHO nd Multipoint Immulite Toxo IgG Immulite 000. WHO rd Two point Modular Toxo IgG Cobas e Cobas e01 Modular E Elecsys WHO rd Two point Downloaded from on August 1, 01 by guest

17 Table. Relative sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) testing of 1 positive and 1 negative non selected samples for anti-toxoplasma Immunoglobulin G (IgG) a. These criteria have been determined with the high and low cut-offs specified by the manufacturers. Confirmatory test : Western Blot LDBIO Toxo II IgG test Sensitivity Specificity PPV NPV (%) a (%) a (%) b (%) b Access Toxo IgG I equivocal result considered negative equivocal result considered positive Access Toxo IgG II equivocal result considered negative equivocal result considered positive Vidas Toxo IgG equivocal result considered negative equivocal result considered positive AxSYM Toxo IgG equivocal result considered negative equivocal result considered positive Immulite Toxo IgG equivocal result considered negative equivocal result considered positive Modular Toxo IgG equivocal result considered negative equivocal result considered positive a Performance values were calculated using Western Blot LDBIO Toxo II IgG test as the reference test. b PPVs and NPVs were calculated using the seroprevalence of our sampling (.1%) which is very close to the French seroprevalence (.%) (1). Downloaded from on August 1, 01 by guest 1

18 Table. Qualitative results of the discordant sera out of 0 with the anti-toxoplasma IgG automated immunoassays and comparison with the Toxo II IgG Western Blot results. Western Blot Total N of Samples Test results for automated immunoassay IgG I Access IgG II Access IgG Axsym IgG Vidas IgG Modular IgG Immulite N a E b P c N a E b P c N a E b P c N a E b P c N a E b P c N a E b P c N a 1 1 E b P c a N negative, b E equivocal, c P positive For the WB, results were considered equivocal when there were only bands one of which being p0. Results of each automated immunoassay and the LDbioToxo II IgG Western Blot (WB) using the cut-offs specified by the manufacturers. Downloaded from on August 1, 01 by guest