Nanoextraction Coupled to Liquid Chromatography Mass Spectrometry Delivers Improved Spatially Resolved Analysis

Abstract

Direct analyte-probed nanoextraction (DAPNe) is a technique that allows extraction of drug and endogenous compounds from a discrete location on a tissue sample using a nano capillary filled with solvent. Samples can be extracted from spot diameters as low as 6 μm. Studies previously undertaken by our group have shown that the technique can provide good precision (5%) for analyzing drug molecules in 150 μm diameter areas of homogenized tissue, provided an internal standard is sprayed on to the tissue prior to analysis. However, without an isotopically labeled standard, the repeatability is poor, even after normalization to the spot area or matrix compounds. By application to tissue homogenates spiked with drug compounds, we can demonstrate that it is possible to significantly improve the repeatability of the technique by incorporating a liquid chromatography separation step. Liquid chromatography is a technique for separating compounds prior to mass spectrometry (LC-MS) which enables separation of isomeric compounds that cannot be discriminated using mass spectrometry alone, as well as reducing matrix interferences. Conventionally, LC-MS is carried out on bulk or homogenized samples, which means analysis is essentially an average of the sample and does not take into account discrete areas. This work opens a new opportunity for spatially resolved liquid chromatography mass spectrometry with precision better than 20%.

Nanocapillary sampling coupled to liquid chromatography mass spectrometry delivers single cell drug measurement and lipid fingerprints

“…41 Our work using nanocapillary sampling was successful at quantifying local drug concentrations in tissue samples. 41,42 We demonstrated that the addition of a chromatography step into the workflow significantly improves measurement precision compared with nanospray ionisation (NSI), with the further advantage of separating lipid classes and allowing the use of automated databases that rely on peak assignment. Here, we advance this work by using nanocapillary sampling coupled to LC-MS to detect and quantify drug molecules (antibiotics) and simultaneously generate a lipid fingerprint from single living cells.…”

Spatially resolved absolute quantitation in thin tissue by mass spectrometry

“…LMJ-SSP, PLESA, and LESA-LC systems can achieve a smaller sampling spot size down to~0.4 mm. In addition, the Swan probe [40] and direct analyte-probed nanoextraction (DAPNe) [41,42] can be coupled to LC separation and allows extraction from2 00-μm and~100-μm areas, respectively, but the true quantitation performance of these system has not been demonstrated. Ultimately, spatial resolution is dependent on the size of the LMJ with the sample surface.…”

Colocation of Lipids, Drugs, and Metal Biomarkers Using Spatially Resolved Lipidomics with Elemental Mapping

“…Unlike mass spectrometry imaging, DAPNe provides the opportunity for chromatographic separation, to separate isobaric compounds and reduce ion suppression effects. 31 …”

“…In the liquid extraction approach known as direct analyte-probed nanoextraction (DAPNe), a nanospray capillary filled with a solvent is directed at an area of interest on a sample, the solvent is pushed onto the surface to dissolve analytes, and these are aspirated into the capillary tip. Unlike mass spectrometry imaging, DAPNe provides the opportunity for chromatographic separation, to separate isobaric compounds and reduce ion suppression effects …”

“…Unlike mass spectrometry imaging, DAPNe provides the opportunity for chromatographic separation, to separate isobaric compounds and reduce ion suppression effects. 31 In this work, we have developed spatially resolved lipidomics using DAPNe. We show how spatially resolved lipidomics, followed by elemental mapping, can be used to correlate metal accumulation with localized lipid profiles in rabbit lung tissues, following tuberculosis infection.…”