{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,27]],"date-time":"2026-09-27T14:45:46Z","timestamp":1790520346235,"version":"4.1.0"},"reference-count":41,"publisher":"Wiley","issue":"6","license":[{"start":{"date-parts":[[2020,4,14]],"date-time":"2020-04-14T00:00:00Z","timestamp":1586822400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003958","name":"Stichting voor de Technische Wetenschappen","doi-asserted-by":"publisher","award":["13382"],"award-info":[{"award-number":["13382"]}],"id":[{"id":"10.13039\/501100003958","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Clin Experimental Allergy"],"published-print":{"date-parts":[[2020,6]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:sec><jats:title>Background<\/jats:title><jats:p>Skin patch testing is still seen as the gold standard for the diagnosis of allergic hypersensitivity. For several metals and for patients with a suspected adverse reaction to their medical device implant material, patch testing can be unreliable. The current alternative to metal allergy patch testing is the in vitro lymphocyte proliferation test (LPT) using tritiated thymidine. This method is well\u2010established but requires handling of radioactive material, often uses heat\u2010inactivated allogenic human pooled serum and cannot determine T cell subsets.<\/jats:p><\/jats:sec><jats:sec><jats:title>Objective<\/jats:title><jats:p>To develop a radioactive free LPT by using carboxyfluorescein succinimidyl ester (CFSE) and to evaluate the influence of serum source (heat\u2010inactivated human pooled serum [HI HPS] vs autologous serum) on the sensitivity and specificity of the nickel\u2010specific LPT.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>Peripheral blood mononuclear cells derived from nickel\u2010allergic patients and healthy controls were collected, labelled with CFSE and cultured in medium containing 10% HI HPS or 10% autologous serum with or without additional T cell skewing cytokine cocktails (Th1: IL\u20107\/IL\u201012, Th2: IL\u20107\/IL\u20104 or Th17: IL\u20107\/IL\u201023\/IL\u20101\u03b2) in the absence or presence of NiSO<jats:sub>4<\/jats:sub>. The stimulation index (SI) was calculated as the ratio of divided cells, that is the percentage of CFSE<jats:sup>low\/neg<\/jats:sup> CD3<jats:sup>+<\/jats:sup>CD4<jats:sup>+<\/jats:sup> T\u2010lymphocytes upon nickel stimulation compared to the percentage of CFSE<jats:sup>low\/neg<\/jats:sup> CD3<jats:sup>+<\/jats:sup>CD4<jats:sup>+<\/jats:sup> T\u2010lymphocytes without antigen. These results were compared with the history of Ni allergy, patch test results and the MELISA test.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>Autologous serum positively influenced Ni\u2010specific proliferation while HI HPS negatively influenced Ni\u2010specific proliferation. The test protocol analysing CD4<jats:sup>+<\/jats:sup> cells and autologous serum without skewing cytokines scored the best diagnostic values (sensitivity 95%; specificity 93%; and overall accuracy 94%) compared to the parallel test using HI HPS (accuracy 60%). Cytokine supplements did not further improve the test protocol which used autologous serum. The protocol using HI HPS could be further improved by addition of the cytokine skewing cocktails.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions<\/jats:title><jats:p>Here, we describe an optimized and highly accurate flow cytometric LPT which comprises of CFSE\u2010labelled cells cultured in autologous serum (not heat inactivated) and without the presence of T cell skewing cytokines.<\/jats:p><\/jats:sec><jats:sec><jats:title>Clinical relevance<\/jats:title><jats:p>The sensitivity and specificity of LPT is enhanced, compared to HI HPS, when autologous serum without skewing cytokines is used.<\/jats:p><\/jats:sec>","DOI":"10.1111\/cea.13603","type":"journal-article","created":{"date-parts":[[2020,3,26]],"date-time":"2020-03-26T07:54:33Z","timestamp":1585209273000},"page":"722-732","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":12,"title":["Non\u2013heat inactivated autologous serum increases accuracy of in vitro CFSE lymphocyte proliferation test (LPT) for nickel"],"prefix":"10.1111","volume":"50","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9170-5302","authenticated-orcid":false,"given":"Niels P. J.","family":"de Graaf","sequence":"first","affiliation":[{"name":"Department of Dermatology Amsterdam UMC Vrije Universiteit Amsterdam Amsterdam The Netherlands"},{"name":"Department of Dental Materials Science Academic Centre for Dentistry Amsterdam University of Amsterdam and Vrije Universiteit Amsterdam Amsterdam The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hetty J.","family":"Bontkes","sequence":"additional","affiliation":[{"name":"Department of Clinical Chemistry Medical Immunology Laboratory Amsterdam Infection &amp; Immunity Amsterdam UMC Vrije Universiteit Amsterdam Amsterdam The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sanne","family":"Roffel","sequence":"additional","affiliation":[{"name":"Department of Oral Cell Biology Academic Centre for Dentistry Amsterdam MOVE Research Institute Amsterdam University of Amsterdam and Vrije Universiteit Amsterdam Amsterdam The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cornelis J.","family":"Kleverlaan","sequence":"additional","affiliation":[{"name":"Department of Dental Materials Science Academic Centre for Dentistry Amsterdam University of Amsterdam and Vrije Universiteit Amsterdam Amsterdam The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Thomas","family":"Rustemeyer","sequence":"additional","affiliation":[{"name":"Department of Dermatology Amsterdam UMC Vrije Universiteit Amsterdam Amsterdam The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sue","family":"Gibbs","sequence":"additional","affiliation":[{"name":"Department of Oral Cell Biology Academic Centre for Dentistry Amsterdam MOVE Research Institute Amsterdam University of Amsterdam and Vrije Universiteit Amsterdam Amsterdam The Netherlands"},{"name":"Department of Molecular Cell Biology and Immunology Amsterdam Infection &amp; Immunity Amsterdam UMC Vrije Universiteit Amsterdam Amsterdam The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Albert J.","family":"Feilzer","sequence":"additional","affiliation":[{"name":"Department of Dental Materials Science Academic Centre for Dentistry Amsterdam University of Amsterdam and Vrije Universiteit Amsterdam Amsterdam The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2020,4,14]]},"reference":[{"key":"e_1_2_7_2_1","doi-asserted-by":"publisher","DOI":"10.12688\/f1000research.13499.1"},{"key":"e_1_2_7_3_1","doi-asserted-by":"publisher","DOI":"10.1111\/cod.12327"},{"key":"e_1_2_7_4_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.0105-1873.2004.00351.x"},{"key":"e_1_2_7_5_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1600-0536.2012.02070.x"},{"key":"e_1_2_7_6_1","doi-asserted-by":"publisher","DOI":"10.1097\/01.all.0000244794.03239.8e"},{"key":"e_1_2_7_7_1","doi-asserted-by":"publisher","DOI":"10.1111\/cod.13010"},{"key":"e_1_2_7_8_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.0906-6705.2005.00344.x"},{"issue":"1","key":"e_1_2_7_9_1","first-page":"17","article-title":"LTT\u2010MELISA is clinically relevant for detecting and monitoring metal sensitivity","volume":"27","author":"Valentine\u2010Thon E","year":"2006","journal-title":"Neuro Endocrinol Lett"},{"key":"e_1_2_7_10_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jaci.2013.03.023"},{"key":"e_1_2_7_11_1","doi-asserted-by":"publisher","DOI":"10.1111\/cod.12751"},{"key":"e_1_2_7_12_1","doi-asserted-by":"publisher","DOI":"10.1002\/cyto.b.20015"},{"key":"e_1_2_7_13_1","doi-asserted-by":"publisher","DOI":"10.1016\/S1074-7613(00)80605-6"},{"key":"e_1_2_7_14_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.95.16.9488"},{"key":"e_1_2_7_15_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jim.2003.09.004"},{"key":"e_1_2_7_16_1","doi-asserted-by":"publisher","DOI":"10.1078\/0171-2985-00196"},{"key":"e_1_2_7_17_1","doi-asserted-by":"publisher","DOI":"10.1111\/all.12553"},{"key":"e_1_2_7_18_1","doi-asserted-by":"publisher","DOI":"10.4049\/jimmunol.0901767"},{"key":"e_1_2_7_19_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jaci.2008.09.036"},{"key":"e_1_2_7_20_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2133.2009.09400.x"},{"key":"e_1_2_7_21_1","doi-asserted-by":"publisher","DOI":"10.1111\/cod.12432"},{"key":"e_1_2_7_22_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2249.2009.04041.x"},{"key":"e_1_2_7_23_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1600-0536.2007.01045.x"},{"key":"e_1_2_7_24_1","doi-asserted-by":"publisher","DOI":"10.1016\/0887-2333(94)90233-X"},{"key":"e_1_2_7_25_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2222.2004.02045.x"},{"key":"e_1_2_7_26_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0889-5406(03)00239-7"},{"key":"e_1_2_7_27_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0022-1759(01)00297-6"},{"issue":"1","key":"e_1_2_7_28_1","first-page":"189","article-title":"Metal\u2010specific lymphocyte reactivity is downregulated after dental metal replacement","volume":"27","author":"Yaqob A","year":"2006","journal-title":"Neuro Endocrinol Lett"},{"key":"e_1_2_7_29_1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0101814"},{"key":"e_1_2_7_30_1","doi-asserted-by":"publisher","DOI":"10.1111\/jdv.14931"},{"key":"e_1_2_7_31_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2222.2008.02970.x"},{"key":"e_1_2_7_32_1","doi-asserted-by":"publisher","DOI":"10.1111\/cod.12043"},{"key":"e_1_2_7_33_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jid.2017.05.025"},{"key":"e_1_2_7_34_1","doi-asserted-by":"publisher","DOI":"10.1111\/cea.13040"},{"key":"e_1_2_7_35_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1523-1747.2003.12623.x"},{"key":"e_1_2_7_36_1","doi-asserted-by":"publisher","DOI":"10.1186\/1479-5876-4-40"},{"key":"e_1_2_7_37_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.immuni.2016.08.003"},{"key":"e_1_2_7_38_1","doi-asserted-by":"publisher","DOI":"10.4049\/jimmunol.1301626"},{"key":"e_1_2_7_39_1","doi-asserted-by":"publisher","DOI":"10.4049\/jimmunol.180.1.510"},{"key":"e_1_2_7_40_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.immuni.2008.02.001"},{"key":"e_1_2_7_41_1","doi-asserted-by":"publisher","DOI":"10.1039\/C8NR07424K"},{"key":"e_1_2_7_42_1","doi-asserted-by":"publisher","DOI":"10.1111\/1523-1747.ep12494841"}],"container-title":["Clinical &amp; 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