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Accurate and sensitive quantification of tacrolimus in small\u2010volume mouse serum is essential not only for pharmacokinetic studies but also for evaluating in\u00a0vivo P\u2010gp functional activity, particularly in pregnancy\u2010related drug transport research. Tacrolimus was extracted using methyl tert\u2010butyl ether after alkalization of the serum with NH\u2084OH. Chromatographic separation was achieved on a C\n                    <jats:sub>18<\/jats:sub>\n                    HPLC column with a mobile phase of acetonitrile and water containing 0.2% NH\u2084OH. Detection was performed in negative ion mode using multiple reaction monitoring (MRM) transitions of\n                    <jats:italic>m<\/jats:italic>\n                    \/\n                    <jats:italic>z<\/jats:italic>\n                    802.5\u2009\u2192\u2009560.6 for tacrolimus and\n                    <jats:italic>m<\/jats:italic>\n                    \/\n                    <jats:italic>z<\/jats:italic>\n                    805.5\u2009\u2192\u2009563.6 for the internal standard, tacrolimus\u2010\n                    <jats:sup>13<\/jats:sup>\n                    <jats:italic>C<\/jats:italic>\n                    ,\n                    <jats:italic>d<\/jats:italic>\n                    <jats:sub>\n                      <jats:italic>2<\/jats:italic>\n                    <\/jats:sub>\n                    . The calibration range for tacrolimus was 0.26\u201344.8\u2009ng\/mL. The method demonstrated good precision, with intra\u2010 and inter\u2010day relative standard deviations below 12%, and accuracy ranging from 94% to 103%. Compared to previously published methods, this approach requires only 200\u2009\u03bcL of mouse serum and provides high sensitivity, with a lower limit of quantification of 0.26\u2009ng\/mL. The assay showed high sensitivity, minimal matrix interference, and good stability across tested conditions.\n                  <\/jats:p>","DOI":"10.1002\/bmc.70437","type":"journal-article","created":{"date-parts":[[2026,4,9]],"date-time":"2026-04-09T01:14:54Z","timestamp":1775697294000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Development and Validation of an LC\u2013MS\/MS Method for Quantitative Determination of Tacrolimus in Mouse Serum"],"prefix":"10.1002","volume":"40","author":[{"given":"Xiaoming","family":"Wang","sequence":"first","affiliation":[{"name":"Mass Spectrometry Core Facility, Division of Basic Science &amp; Transitional Research, Department of Obstetrics &amp; Gynecology University of Texas Medical Branch  Galveston Texas 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