The March of the Masses

Abstract

The growth of mass spectrometry and more specifically liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) 2 continues to accelerate in the clinical laboratory. A recent publication (1 ) provides readers with the opportunity to see where we have been, offering examples of how LC-MS/MS has impacted diagnostic care (steroids and clinical toxicology); where we are (pain management, peptide hormones, and single proteins); and where we are going (protein panels, increased throughput and automation). The techniques and technologies discussed posit 3 conclusions. First, the potential scope of mass spectrometry testing in clinical chemistry is prospectively limitless. Second, the tools and technologies being deployed will require further refinement for mass spectrometers to become established automated clinical analyzers. Third, the promise of reference interval measurement in support of clinical measurement has been practically realized for some applications and is feasible for others.Two central tenets are evident in each application. First, the "killer app" of mass spectrometry incorporates the use of a stable labeled isotope of the analyte(s) being measured with various isotope dilution strategies for calibration. An ideal internal standard is both physiologically unique relative to the composition of the test specimen and physicochemically identical to the analyte, such that recovery, separation, and ionization in each specimen are controlled and corrected: the perfect analytical control for each specimen. Second, multiconcentration calibrators defining the analytical range, often bracketing QCs and samples, control for instrument response drift and deviations from detection linearity over the course of analysis. The combination of these two principles, coupled with the specificity of measurement that LC-MS/MS affords, provides the opportunity for reference level measurement. This, however, comes with historical baggage; LC-MS/MS is almost exclusively deployed in a batch mode of analysis in clinical laboratories.In this issue of Clinical Chemistry, Olson et al. (2 ) illuminate, illustrate, and iterate the viewpoint that the dogma of batch mode analysis is both outdated and unnecessarily burdensome. The practice of calibration curve analysis with each set of samples is derived from pharmaceutical drug development from the mid1990s. Application of LC-MS/MS, the gold standard technique, was hampered by the technologies of the time and adherence to guidance, consequently resulting in ultraconservative analytical methodologies (3 ). The constraint of daily or batch-specific calibration is most keenly felt in the clinical laboratory, particularly in smaller hospitals settings where STAT sample analysis is de rigueur and a single instrument is required to perform many different measurements on a diverse array of analytes within a single shift. The obvious need for random access LC-MS/MS systems is apparent (1 ), and Olson et al. provide insights and key details in the path toward this goal. Most importan...

Analytical Performance of LC-MS/MS Method for Simultaneous Determination of Five Steroids in Serum

“…Other authors achieve similar results to our method using online extraction and APCI ionization, and silica column for steroid separation, however need at least 600 µL of serum sample and longer analysis time [6,12]. Other methods used manual sample preparation employing solid-phase or liquid-liquid extraction, evaporation or/and derivatization steps and although adequate for steroid analysis, total Functional sensitivity (ng/mL) 0.5 0.25 0.5 0.5 0.5 …”

“…The last generation of tandem mass spectrometers has superior limits of quantification, permitting omission or previously employed derivatization steps [11]. In the present study we evaluated a method to quantify adrenal steroids by liquid chromatography tandem mass spectrometry (LC-MS/MS) We developed a method to measure five steroids using protein precipitation, isotopic internal standards and two dimensional liquid chromatography, consisting of trapping column and reverse-phase C18 core-shell analytical column following atmospheric pressure chemical ionization and mass spectrometry detection with a total run time of 6.4 minutes including on-line extraction [6]. The combination of on-line extraction and the use of the corresponding deuterated internal standards reduced variability and contributed to the good precision achieved for all compounds.…”

“…Tandem mass spectrometry methods with atmospheric pressure chemical ionization (APCI) ionization were developed in 1990s [4] and during the past two decades, LC-MS/MS techniques showed an increased usage in clinical laboratories, mainly to small molecules analyses [5,6]. The accurate measurement of hormones is the pivot of modern endocrinology and so lays the importance of method validation to demonstrate characteristics of reprodutibility, accuracy and specificity [7,8].…”

Alternative for Reducing Calibration Standard Use in Mass Spectrometry

“…Recent publications have discussed alternative calibration strategies for quantitative clinical mass spectrometry and proposed terminology for these approaches (1)(2)(3). Like Grant (3 ), we believe that mass spectrometry laboratories generally use more calibration standards than necessary.…”

Diagnostic application of the exponentially modified Gaussian model for peak quality and quantitation in high-throughput liquid chromatography–tandem mass spectrometry

“…Calculating the chromatographic peak area in the absence of noise is a straightforward numerical integration process of peak detection, peak boundary assignment, and baseline estimation [1]. The move to apply liquid chromatography-tandem mass spectrometry (LC-MS/MS) in this setting has revolutionized accuracy, precision and selectivity across a wide range of target molecules [2][3][4][5][6] and allowed for the use of smaller sample volumes while achieving lower detection limits. Biological analytes have benefited from atmospheric pressure ionization techniques, but the drive to report at the detection limit necessarily produces results that are significantly more noise-prone than systems investigated by the older electron impact methods.…”