{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,12,23]],"date-time":"2025-12-23T18:46:11Z","timestamp":1766515571077,"version":"3.40.5"},"reference-count":28,"publisher":"Public Library of Science (PLoS)","issue":"9","license":[{"start":{"date-parts":[[2022,9,14]],"date-time":"2022-09-14T00:00:00Z","timestamp":1663113600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Health System Research Institute","award":["HSRI 63-079"],"award-info":[{"award-number":["HSRI 63-079"]}]},{"DOI":"10.13039\/501100004704","name":"National Research Council of Thailand","doi-asserted-by":"crossref","award":["GSCMU (NRCT)\/15\/2563"],"award-info":[{"award-number":["GSCMU (NRCT)\/15\/2563"]}],"id":[{"id":"10.13039\/501100004704","id-type":"DOI","asserted-by":"crossref"}]},{"name":"Chiang Mai University Grant for New Researcher"}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS ONE"],"abstract":"<jats:p>Mutations of the n<jats:italic>ucleophosmin1<\/jats:italic> (<jats:italic>NPM1<\/jats:italic>) gene represent the most frequent molecular alteration in acute myelogenous leukemia (AML), especially in patients with AML who have a normal karyotype. These alterations have been shown to carry favorable prognostic significance in patients with AML. Several methods have been developed for detection of <jats:italic>NPM1<\/jats:italic> gene mutations. However, their ability to detect low levels of mutations in a wild-type background is limited. In this study, the Enhance improved and complete enrichment Co-amplification at Lower Denaturation temperature Polymerase Chain Reaction (E-ice-COLD-PCR) assay combined with High Resolution Melting (HRM) analysis was developed and validated for highly specific and sensitive screening for <jats:italic>NPM1<\/jats:italic> gene mutations. A total of 83 blood samples from patients with AML were collected, and their DNA was extracted. For mutational analysis, the E-ice-COLD-PCR assay for the detection of <jats:italic>NPM1<\/jats:italic> gene mutations was developed. PCR products were analyzed by HRM analysis. All positive samples were confirmed by direct sequencing. This assay enabled detection specificity and sensitivity of <jats:italic>NPM1<\/jats:italic> mutations in 9\/83 patients with AML. Direct sequencing results were 100% concordant with this method. In addition, the limit of detection was 12.5% mutant in the final concentration of 5 ng genomic DNA. The E-ice-COLD-PCR assay with HRM analysis is a highly specific and sensitive screening method for enrichment of detecting <jats:italic>NPM1<\/jats:italic> gene mutations. This method has both a short turn around time and easier interpretation compared to those of other methods.<\/jats:p>","DOI":"10.1371\/journal.pone.0274034","type":"journal-article","created":{"date-parts":[[2022,9,14]],"date-time":"2022-09-14T17:50:55Z","timestamp":1663177855000},"page":"e0274034","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":2,"title":["Development of E-ice-COLD-PCR assay combined with HRM analysis for Nucleophosmin1 gene mutation detection in acute myelogenous leukemia"],"prefix":"10.1371","volume":"17","author":[{"given":"Rattana","family":"Kongta","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Noppamas","family":"Panyasit","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Wuttichote","family":"Jansaento","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8436-1695","authenticated-orcid":true,"given":"Suwit","family":"Duangmano","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"340","published-online":{"date-parts":[[2022,9,14]]},"reference":[{"volume-title":"Cancer in Thailand","year":"2021","author":"J Rojanamatin","key":"pone.0274034.ref001"},{"key":"pone.0274034.ref002","doi-asserted-by":"crossref","first-page":"2040620719899818","DOI":"10.1177\/2040620719899818","article-title":"Nucleophosmin1 (NPM1) abnormality in hematologic malignancies, and therapeutic targeting of mutant NPM1 in acute myeloid leukemia","volume":"11","author":"Y Chen","year":"2020","journal-title":"Ther Adv Hematol"},{"issue":"1","key":"pone.0274034.ref003","article-title":"The Role of Nucleophosmin 1 (NPM1) Mutation in the Diagnosis and Management of Myeloid Neoplasms","volume":"12","author":"K. 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