No abstract

A novel single cell analytical method showing adipocyte heterogeneity dependent on lipid droplet size using nanomanipulation-coupled nanoelectrospray mass spectrometry.

Electrospray mass spectrometry is now a widely used technique for observing cell content of various biological tissues. However, electrospray techniques (liquid chromatography and direct infusion) often involve lysing a group of cells and extracting the biomolecules of interest, rather than a sensitive, individual cell method to observe local chemistry. Presented here is an approach of combining a nanomanipulator workstation with nanospray mass spectrometry, which allows for extraction of a single cell, followed by rapid mass analysis that can provide a detailed metabolic profile. Triacylglycerol content was profiled with this tool coupled to mass spectrometry to investigate heterogeneity between healthy and tumorous tissues as well as lipid droplet containing adipocytes in vitro as proof of concept. This selective approach provides cellular resolution and complements existing bioanalytical techniques with minimal invasion to samples. In addition, the coupling of nanomanipulation and mass spectrometry holds the potential to be used in a great number of applications for individual organelles, diseased tissues, and in vitro cell cultures for observing heterogeneity even amongst cells and organelles of the same tissue.

Abstract. We describe a novel technique combining precise organelle microextraction with deposition and matrix-assisted laser desorption/ionization (MALDI) for a rapid, minimally invasive mass spectrometry (MS) analysis of single organelles from living cells. A dual-positioner nanomanipulator workstation was utilized for both extraction of organelle content and precise co-deposition of analyte and matrix solution for MALDI-direct organelle mass spectrometry (DOMS) analysis. Here, the triacylglycerol (TAG) profiles of single lipid droplets from 3T3-L1 adipocytes were acquired and results validated with nanoelectrospray ionization (NSI) MS. The results demonstrate the utility of the MALDI-DOMS technique as it enabled longer mass analysis time, higher ionization efficiency, MS imaging of the co-deposited spot, and subsequent MS/MS capabilities of localized lipid content in comparison to NSI-DOMS. This method provides selective organellar resolution, which complements current biochemical analyses and prompts for subsequent subcellular studies to be performed where limited samples and analyte volume are of concern.