Predicted influence of sample hematocrit on injected mass of internal standard in mass spectrometry assays utilizing simple protein precipitation for sample preparation

Allison M Cushman-Vokoun, Kim B. Plath, James D. Landmark, Douglas F. Stickle

Research output: Contribution to journalArticle

Abstract

Background: The injected mass of internal standard is often monitored in mass spectrometry assays to flag aberrant specimen processing. When simple protein precipitation is used for sample preparation, the protein volume (solids fraction) of the specimen is in principle a variable affecting the final concentration of the internal standard in the liquid phase. We examined the predicted extent of this variation for an example of protein precipitation for sample preparation used in a mass spectrometry assay for immunosuppressants in whole blood. Methods: Liquid and solid mass fractions of samples with hematocrit ranging from 15% to 60% were measured after protein precipitation of samples using a whole blood/precipitating reagent ratio of 1:4. Relative supernatant volumes as a function of hematocrit were determined. Results: Liquid volume variation was consistent with a predicted variation in the internal standard concentration of only approximately ±1% across this hematocrit range. Data were in close agreement with calculations based simply on protein density and concentration. Conclusions: Hematocrit affects the injected mass of internal standard in assays that utilize protein precipitation for sample preparation, due to predictable variation in solids and liquid fractions. The effect is small, however, compared to the typical variation observed for injected mass of internal standard.

Original languageEnglish (US)
Pages (from-to)215-219
Number of pages5
JournalClinical Chemistry and Laboratory Medicine
Volume45
Issue number2
DOIs
StatePublished - Feb 1 2007

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Hematocrit
Mass spectrometry
Assays
Mass Spectrometry
Proteins
Liquids
Blood
Immunosuppressive Agents
Processing

Keywords

  • Hematocrit
  • Internal standard
  • Liquid chromatography-tandem mass spectrometry
  • Quality control

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Biochemistry, medical

Cite this

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title = "Predicted influence of sample hematocrit on injected mass of internal standard in mass spectrometry assays utilizing simple protein precipitation for sample preparation",
abstract = "Background: The injected mass of internal standard is often monitored in mass spectrometry assays to flag aberrant specimen processing. When simple protein precipitation is used for sample preparation, the protein volume (solids fraction) of the specimen is in principle a variable affecting the final concentration of the internal standard in the liquid phase. We examined the predicted extent of this variation for an example of protein precipitation for sample preparation used in a mass spectrometry assay for immunosuppressants in whole blood. Methods: Liquid and solid mass fractions of samples with hematocrit ranging from 15{\%} to 60{\%} were measured after protein precipitation of samples using a whole blood/precipitating reagent ratio of 1:4. Relative supernatant volumes as a function of hematocrit were determined. Results: Liquid volume variation was consistent with a predicted variation in the internal standard concentration of only approximately ±1{\%} across this hematocrit range. Data were in close agreement with calculations based simply on protein density and concentration. Conclusions: Hematocrit affects the injected mass of internal standard in assays that utilize protein precipitation for sample preparation, due to predictable variation in solids and liquid fractions. The effect is small, however, compared to the typical variation observed for injected mass of internal standard.",
keywords = "Hematocrit, Internal standard, Liquid chromatography-tandem mass spectrometry, Quality control",
author = "Cushman-Vokoun, {Allison M} and Plath, {Kim B.} and Landmark, {James D.} and Stickle, {Douglas F.}",
year = "2007",
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AU - Landmark, James D.

AU - Stickle, Douglas F.

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N2 - Background: The injected mass of internal standard is often monitored in mass spectrometry assays to flag aberrant specimen processing. When simple protein precipitation is used for sample preparation, the protein volume (solids fraction) of the specimen is in principle a variable affecting the final concentration of the internal standard in the liquid phase. We examined the predicted extent of this variation for an example of protein precipitation for sample preparation used in a mass spectrometry assay for immunosuppressants in whole blood. Methods: Liquid and solid mass fractions of samples with hematocrit ranging from 15% to 60% were measured after protein precipitation of samples using a whole blood/precipitating reagent ratio of 1:4. Relative supernatant volumes as a function of hematocrit were determined. Results: Liquid volume variation was consistent with a predicted variation in the internal standard concentration of only approximately ±1% across this hematocrit range. Data were in close agreement with calculations based simply on protein density and concentration. Conclusions: Hematocrit affects the injected mass of internal standard in assays that utilize protein precipitation for sample preparation, due to predictable variation in solids and liquid fractions. The effect is small, however, compared to the typical variation observed for injected mass of internal standard.

AB - Background: The injected mass of internal standard is often monitored in mass spectrometry assays to flag aberrant specimen processing. When simple protein precipitation is used for sample preparation, the protein volume (solids fraction) of the specimen is in principle a variable affecting the final concentration of the internal standard in the liquid phase. We examined the predicted extent of this variation for an example of protein precipitation for sample preparation used in a mass spectrometry assay for immunosuppressants in whole blood. Methods: Liquid and solid mass fractions of samples with hematocrit ranging from 15% to 60% were measured after protein precipitation of samples using a whole blood/precipitating reagent ratio of 1:4. Relative supernatant volumes as a function of hematocrit were determined. Results: Liquid volume variation was consistent with a predicted variation in the internal standard concentration of only approximately ±1% across this hematocrit range. Data were in close agreement with calculations based simply on protein density and concentration. Conclusions: Hematocrit affects the injected mass of internal standard in assays that utilize protein precipitation for sample preparation, due to predictable variation in solids and liquid fractions. The effect is small, however, compared to the typical variation observed for injected mass of internal standard.

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KW - Quality control

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