TY - JOUR
T1 - The suitability of matrix assisted laser desorption/ionization time of flight mass spectrometry in a laboratory developed test using cystic fibrosis carrier screening as a model
AU - Farkas, Daniel H.
AU - Miltgen, Nicholas E.
AU - Stoerker, Jay
AU - Van Den Boom, Dirk
AU - Highsmith, W. Edward
AU - Cagasan, Lesley
AU - McCullough, Ron
AU - Mueller, Reinhold
AU - Tang, Lin
AU - Tynan, John
AU - Tate, Courtney
AU - Bombard, Allan
PY - 2010/9
Y1 - 2010/9
N2 - We designed a laboratory developed test (LDT) by using an open platform for mutation/polymorphism detection. Using a 108-member (mutation plus variant) cystic fibrosis carrier screening panel as a model, we completed the last phase of LDT validation by using matrix-assisted laser desorption/ionization time of flight mass spectrometry. Panel customization was accomplished via specific amplification primer and extension probe design. Amplified genomic DNA was subjected to allele specific, single base extension endpoint analysis by mass spectrometry for inspection of the cystic fibrosis transmembrane regulator gene (NM-000492.3). The panel of mutations and variants was tested against 386 blinded samples supplied by "authority" laboratories highly experienced in cystic fibrosis transmembrane regulator genotyping; >98% concordance was observed. All discrepant and discordant results were resolved satisfactorily. Taken together, these results describe the concluding portion of the LDT validation process and the use of mass spectrometry to detect a large number of complex reactions within a single run as well as its suitability as a platform appropriate for interrogation of scores to hundreds of targets.
AB - We designed a laboratory developed test (LDT) by using an open platform for mutation/polymorphism detection. Using a 108-member (mutation plus variant) cystic fibrosis carrier screening panel as a model, we completed the last phase of LDT validation by using matrix-assisted laser desorption/ionization time of flight mass spectrometry. Panel customization was accomplished via specific amplification primer and extension probe design. Amplified genomic DNA was subjected to allele specific, single base extension endpoint analysis by mass spectrometry for inspection of the cystic fibrosis transmembrane regulator gene (NM-000492.3). The panel of mutations and variants was tested against 386 blinded samples supplied by "authority" laboratories highly experienced in cystic fibrosis transmembrane regulator genotyping; >98% concordance was observed. All discrepant and discordant results were resolved satisfactorily. Taken together, these results describe the concluding portion of the LDT validation process and the use of mass spectrometry to detect a large number of complex reactions within a single run as well as its suitability as a platform appropriate for interrogation of scores to hundreds of targets.
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U2 - 10.2353/jmoldx.2010.090233
DO - 10.2353/jmoldx.2010.090233
M3 - Article
C2 - 20616359
AN - SCOPUS:77956819100
SN - 1525-1578
VL - 12
SP - 611
EP - 619
JO - Journal of Molecular Diagnostics
JF - Journal of Molecular Diagnostics
IS - 5
ER -