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Application of Various Normalization Methods for Microscale Analysis of Tissues Using Direct Analyte Probed

Janella de Jesus1,2, Josephine Bunch2, Guido Verbeck3

  • 1Department of Chemistry , University of Surrey , Guildford , Surrey GU2 7XH , United Kingdom.

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Direct analyte probed nanoextraction (DAPNe) improves mass spectrometry sensitivity. This study enhances DAPNe

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Area of Science:

  • Analytical Chemistry
  • Mass Spectrometry
  • Nanotechnology

Background:

  • Direct analyte probed nanoextraction (DAPNe) offers high sensitivity for microscale sample analysis.
  • Previous applications include single-cell and organelle analysis.
  • Quantitative capabilities of DAPNe require further evaluation.

Purpose of the Study:

  • To investigate normalization techniques for improving DAPNe quantitative accuracy.
  • To assess methods for internal standard incorporation.
  • To explore normalization strategies when internal standards are unavailable.

Main Methods:

  • Investigated two internal standard incorporation methods on test substrates mimicking biological matrices.
  • Evaluated normalization to extraction spot area.
  • Assessed normalization to matrix compounds.

Main Results:

  • Sprayed internal standards significantly reduced observed variability.
  • Normalization to the extracted area also reduced variability in certain scenarios.
  • The study identified effective strategies for enhancing DAPNe quantification.

Conclusions:

  • Normalization techniques are crucial for improving the quantitative performance of DAPNe.
  • Internal standards, particularly sprayed ones, are effective for reducing variability.
  • Matrix-based normalization offers an alternative when internal standards are not feasible.