(A) Chemical structure and predicted molecular weight of methylamine adduct of PTAD-derivatized 24,25(OH)2D3. (B) MS/MS fragmentation spectra of methylamine adduct of PTAD-derivatized 24,25(OH)2D3. (C) LC-MRM elution peaks of 24,25(OH)2D3 and 25D3 metabolites and their respective isotopic standards from a representative patient sample run. Mass transitions are shown. (D) Assay linearity of serially diluted samples, n=4 replicates per dilution, standard error bars shown. (E) Coefficients of variance of sample assay measurements plotted against analyte concentration; n=4 replicate measurements per data point.