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Amanitin ELISA Management of suspected poisoning

The only commercial Amanitin ELISA In Case of Emergency: Results within 1 hour

Accurate

Highly Sensitive

Excellent Negative Predictive Value

CF032-03E Amanitin ELISA

Assay Performance Data Epidemiology ELISA Intra-assay precision 6.3% intoxication has been reported Precoated Microtiter Plate (n=4 urine samples, range: 3.1-90.8 ng/ml; to be lethal in up to 25 % of the cases. wash 2x 20 duplicates, CV range: 2.8-14%) Mortality rate in children under 10 years Inter-assay precision 7.3% of age is above 50 %. 50 µl Sample (diluted 1:25), (n=3 urine samples; range 6.7-81,8 ng/ml; Clinical Effects and Laboratory Analysis Standard or Control+ 20 runs; CV range: 5.3-10.6%) infiltrate hepatocytes in which 50 µl Amanitin-Biotin Conjugate Dilution linearity 97.8% their primary action is to inhibit nucleo- (3 urine samples diluted 1:25, 50, 100, incubate 30 min at RT (18-28°C) plasmic RNA polymerase II (which in turn 200, 400 & 800; Range: 79.0-119.0%) on a plate rotator interferes with mRNA synthesis). This wash 3 x Spiking recovery 99.9% results in the arrest of protein synthesis (n=1 urine sample 4 times spiked and and cellular necrosis ultimately leading to add 100 µl of Enzyme Label tested; Range: 80.0-127.5%) severe acute hepatitis. Amanitin also acts incubate 15 min at RT (18-28°C) Analytical sensitivity 0.22 ng/ml as a direct nephrotoxin. The symptoms of on a plate rotator (n=20, measured in duplicates) phallatoxic Amanita poisoning are of delayed onset type. Patients may there- wash 3 x Functional sensitivity 1.5 ng/ml fore not primarily associate their symptoms (cut-off: intra-assay CV = 15%) with the ingestion of the wild mushroom. add 100 µl TMB Substrate Specificity Once symptoms appear, amatoxins can incubate 15 min at RT (18-28°C) a-Amanitin: 100% e-Amanitin: 0.1% be detected in urine. The detection of a- on a plate rotator b-Amanitin: 0.1% Phalloidine: not detect. Amanitin (100 %) and g-Amanitin (90 %) g-Amanitin: 90% Phallacidine: not detect. in urine by means of the BÜHLMANN Ama- nitin ELISA confirms Amanita Intoxication. add 100 µl Stop Solution Sample type Urine, Serum, Plasma Amatoxins are detected in urine samples Sample storage and stability within 6-60 hours after ingestion, i.e. the Read at 450 nm 2-8°C: up to 7 days time by which clinical symptoms occur. Time to Result: 1 Hour -20°C: for at least 6 months From internal evaluation data and clinical Standard Range 1-100 ng/ml toxicologic study, capability characteristics may be outlined as follows:

α-Amanitin 1. The Amanitin ELISA test is an additi- Diagnostic accuracy of Amanitin a-Amanitin is the major toxin of the extre- onal tool in an entire diagnostic work- analysis in Urine* up. mely poisonous toadstools, Amanita phal- Diagnostic SENS SPEZ PPV NPV loides (death cap), A. verna (white death 2. A negative result does not exclude a Accuracy (%) cap), A. virosa () and possible Amanitin intoxication. All patients included in the study (n=61) other Amanita species. These Urinary ama- 70 82.4 43.8 93.3 3. Sample collection must be within 36 may also contain other amatoxins such as nitin levels hours.* ≥ 1.5 ng/ml b- and g-Amanitin as well as . Urinary ama- 60 100 100 92.7 Amatoxins are also found in Galerina mar- 4. The Amanitin ELISA test detects a- nitin levels ginata (marginate pholiota), G. autumna- Amanitin (100%) and a-Amanitin ≥ 5.0 ng/ml lis, G. sulciceps and other Galerina and Le- (90%). There is no crossreactivity to Patients evaluated within 36 hours (n=51) piota species. A. phalloides, A. verna and b-Amanitin. Urinary ama- 100 87.2 50 100 A. virosa account for over 90 % of clinical- nitin levels ≥ 1.5 ng/ml ly relevant mushroom intoxication cases in Urinary ama- 100 100 100 100 & Northern America. Amanita spe- *F. Butera et. al: Diagnostic Accuracy of Urinary Amanitin in Suspected : A pilot nitin levels cies also occur in Asia and Australia. Study. J. of Toxicology 2004 ≥ 5.0 ng/ml

BÜHLMANN Laboratories AG Phone +41 61 487 12 12 Baselstrasse 55 Fax orders +41 61 487 12 99 Ordering Code: CH–4124 Schönenbuch/Basel [email protected] EK-AM1 96 wells Switzerland www.buhlmannlabs.ch