26. A Fully-Automated Two Dimensional Liquid Chromatography Isotope Dilution Mass Spectrometry Method for Quantification of Serum Creatinine
Tue 4:42 PM - PosterSplash Track 3
Robert Harlan
Johns Hopkins University
Robert Harlan, William Clarke, Joely Straseski, Danni Meany.

The Johns Hopkins University Department of Pathology, Clinical Chemistry, Baltimore, Maryland
There remains a need for a fast and accurate method of quantitation for creatinine in serum that is both automated and cost effective. Here we demonstrate a fully automated LC-IDMS method for creatinine.

Human serum and an internal standard deuterated creatinine (creatinine D-3) mixture were injected directly onto a Thermo Fisher Cyclone MCX column (0.5 x 50 mm) using the TurboFlow® Technology. This step serves an online solid phase extraction (SPE) to remove big biomolecules such as carbohydrates and phospholipids in the serum and retain the analytes. Then creatinine and creatinine D-3 were eluted to a Thermo Fisher Hypercarb (50 x 3 mm, particle size 5 um) column using a mixture of mobile A (water with 0.1% ammonium hydroxide) and methanol with 2% ammonium hydroxide (80:20, v:v) and separated using a 5-95% gradient of mobile A and mobile B(methanol with 0.1% ammonium hydroxide in 6 minutes. Creatinine and creatinine D3 were ionized by electrospray ionization and measured by their parent ions of m/z 114 and 117,respectively. Precision, linearity, and limits of detection and quantitation of this LC-IDMS method were evaluated.

Inter-day and intra-day coefficients of variation were ≤ 6%. The limit of detection was 0.05 mg/dL and the limit quantitation was 0.3 mg/dL. Average recovery was 104.3% in the range of 0.43 -6.9 mg/dL. Slope for method comparison was 0.96 between this LC-IDMS method and Roche Creatinine Plus enzymatic creatinine assay.

We demonstrate a rapid, fully automated method for quantitative measurement of creatinine in serum.

Support: From Johns Hopkins University
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