Lab Matters Summer 2018 | Page 68

APHL 2018 Annual Meeting Poster Abstracts

Infectious Disease

Background: Routine detection of Measles, Mumps, Rubella and Varicella Zoster( MMRV) IgG is used to determine antibody status where infection history or previous immunization is unknown.

Materials / Methods: This MMRV assay was developed using the Dynex Technologies Multiplier system and coated bead technology. Antigen coated beads representing each MMRV specificity were embedded into the base of carrier 96 well assay plate. Each assay well contains the 4 MMRV targets for the test sample IgG detection. The final chemiluminescent reaction is imaged with the on-board camera and results output as index values referenced against the assay specific calibrator. Precision was measured by assaying a range of 14 samples 3 times across an assay plate on three instruments over three days. A ROC analysis was run in order to set the cut-off for each of MMV and confirm it for Rubella where the cutoff was ultimately defined by the International reference RUBI-1-94. Based on the resulting cut-off values, concordance was assessed on up to 929 samples collected for MMRV screening; results were compared to 510k cleared ELISA assays.

Results: Percentage coefficient of variation(% CV) for each MMRV specificity was calculated and is summarized below:

Precision test Measles(% CV) Mumps(% CV) Rubella(% CV) VZV(% CV) Within run 4.54 5.35 3.69 4.08 Between run 4.51 5.66 4.64 4.06 Between day 3.56 4.01 4.76 3.15 Between instrument 1.41 3.19 2.24 1.37

ROC analysis:

Area under the curve( AUC) and 95 % confidence interval( CI) results were:

• Measles 0.995 AUC( 0.991-0.998 CI)

• Mumps 0.987 AUC( 0.977-0.997CI)

• Rubella 0.998( 0.997-0.999 CI)

• VZV 0.999( 0.997-1.000 CI).

• Percent positive agreement( PPA) and percent negative agreement( PNA) with 95 % confidence intervals( Cl) were calculated in two ways:

• Equivocal samples scored as positive PPA: Measles- 95.3 %( 93.5-96.6 %), Mumps: 90.2( 87.8-92.2 %), Rubella: 93.9( 91.9-95.4 %), VZV: 98.1( 96.8-98.8 %). PNA: Measles – 94.2( 90.7-97.0 %), Mumps: 93.3( 88.5-96.2 %), Rubella 99.5 %( 97.4- 99.9 %), VZV 97.5( 96.3-99.2 %).

• Equivocal samples scored as negative PPA: Measles – 93.3 %( 91.2-95.0 %) Mumps: 93.3( 91.1-95.0 %), Rubella: 93.0( 90.9- 94.7 %), VZV: 97.7( 96.3-98.5 %). PNA: Measles – 95.4( 92.0- 97.4 %), Mumps: 94.6( 90.8-96.9 %), Rubella: 100.0( 98.4- 100.00 %), VZV: 99.2( 95.6-99.9 %).

Conclusion: This multiplexed fully automated assay gives reproducible semi-quantitative results for MMRV IgG. It is ideal for batch testing as can handle up to ninety two test samples in a single plate to produce 368 results in < 3 hours. When two plates are run together 736 results are generated in five hours.

Presenter: Robert Wolfert, PhD, Dynex Technologies, Inc., Chantilly, VA, Phone: 703.803.1254, Email: rwolfert @ dynex. com

Novel ELISA Based on Antigens from Strongyloides papillosus Instead of Strongyloides ratti Exhibits Increased Serological Specificity

B. Menge 1, O. Klemens 1, A. Streit 2, O. Sendscheid 3, J. Klemens 1, K. Steinhagen 1; 1 EUROIMMUN, Lubeck, Germany, 2 Max Planck Institute for Developmental Biology, Tubingen, Germany, 3 EUROIMMUN US, Inc., Mountain Lakes, NJ

Background: Strongyloidiasis is an infectious disease caused by the nematode Strongyloides. Human infection by Strongyloides stercoralis can manifest with dermatological, intestinal and pulmonal symptoms frequently passing into a chronic disease. Low parasitic loads and discontinuous larvae excretion may hamper diagnosis by coproscopy. Serological test systems are more sensitive to detect the infection. Available serological tests are commonly based on native antigens from S. ratti larvae and lack specificity. We developed and evaluated the first ELISA based on S. papillosus to increase specificity. Methods: Evaluation of the ELISA based on S. papillosus was performed using the following three approaches:[ 1 ] Participation in an external quality assessment scheme( NEQAS, UK) encompassing six positive and five negative samples [ 2 ] A correlation study with the commercial Bordier ELISA( Strongyloides ELISA kit based on S. ratti antigens; Bordier Affinity Products, Switzerland) including 89 sera pre-characterized as either positive( n = 59) or negative( n = 30) by means of Bordier ELISA [ 3 ] Comparison with an in house ELISA based on S. ratti by determining specificity with respect to a cross-reactivity panel( n = 193, samples from patients with other parasitic or bacterial infections) and a control panel( n = 688, samples from 500 healthy blood donors, 100 pregnant women and 88 children).

Results: [ 1 ] Results obtained with the Anti-Strongyloides ELISA were 100 % in agreement with NEQAS target values.[ 2 ] In 74 of 89 samples( 83.1 %), the result of the novel ELISA correlated with the Bordier ELISA. Seven discrepant cases, which were positive in Bordier ELISA but negative in the novel ELISA, were further examined. Serological analyses indicated the presence of antibodies against other parasites( Plasmodium spp., Schistosoma spp. and Echinococcus spp.) in six of these cases.[ 3 ] The S. ratti based ELISA was reactive in 13.9 % of the sera in the cross-reactivity panel and in 10.6 % of the samples from healthy individuals, yielding a combined specificity of 88.6 %. In comparison, reactivities of 6.2 %( cross-reactivity panel) and 3.5 %( healthy individuals) were detected with the novel Anti-Strongyloides ELISA, resulting in a combined specificity of 95.9 %.

Discussion: The novel Anti-Strongyloides ELISA reveals a high diagnostic accuracy in the serological diagnosis of Strongyloidiasis. The use of native antigens from S. papillosus instead of S. ratti increases assay specificity by 7.3 %.

Presenter: Oliver Sendscheid, PhD, EUROIMMUN US, Inc., Mountain Lakes, NJ, Phone: 973.656.1000 x130, Email: oliver. sendscheid @ euroimmun. us

Development of a Novel NS1-based ELISA for Detection of Specific IgG Antibodies Against West-Nile Virus

O. Klemens 1, C. Pannwitt 1, A. Hachid 2, J. Klemens 1, O. Sendscheid 3, J. Fraune 1, W. Schlumberger 1, K. Steinhagen 1; 1 EUROIMMUN, Lubeck, Germany, 2 Institut Pasteur d’ Algérie, Dely-Brahim, Algeria

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EUROIMMUN US, Inc., Mountain Lakes, NJ