SpectraX: A Straightforward Tool for Principal Component Analysis-Based Spectral Analysis.

The analysis of complex spectra is an important component of direct/ambient mass spectrometry (MS) applications such as natural product screening. Unlike chromatography-based metabolomics or proteomics approaches, which rely on software and algorithms, the work of spectral screening is mostly performed manually in the initial stages of research and relies heavily on the experience of the analyst. As a result, throughput and spectral screening reliability are problematic when dealing with large amounts of data. Here, we present SpectraX, a MATLAB-based application, which can analyze MS spectra and quickly locate m/z features from them. Principal component analysis (PCA) is used to analyze the data set, and scoring plots are presented to help in understanding the clustering of data. The algorithm uses mass to charge (m/z) features to produce a list of potential natural products.

Micro "Hyper-Channels" on Laser-Refined Cellulose Structures.

Controlled liquid transportation is widely applied in both academia and industry. However, liquid transport applications are limited by parameters such as driving forces, precision, and velocity. Herein, a simple laser-refining technology is presented to produce micro "hyper-channels". A cellulose substrate is rendered hydrophobic through silanization and refined with a laser to produce both hierarchical nanostructures and a wettability contrast simultaneously. Such a method enables faster ("hyper"-channel) aqueous liquid transportation (≈25X, 50 mm s-1 ) compared to conventional methods. Complex patterns can be readily produced at different scales with spatial resolution as low as 50 µm. This technique also controls the refining depth on the thin paper substrate. Shallow channels can be fabricated on thin paper substrates that enable fluidic channel-crossover without liquid mixing. With certain parameters, the technique creates "portals" through the substrate, allowing trans-dimensional liquid transportation between two layers of a single sheet of substrate. The fluid throughput can be increased, while also permitting fluidic channel crossover without liquid mixing. By introducing multiple portals, the controlled fluid can transfer trans-dimensionally several times, enabling further fluidic complexity. The real-life utility of the method is demonstrated by creating a trans-dimensional microfluidic device for colorimetric detection.

Open Port Interface for Coupling Capillary Electrophoresis and Mass Spectrometry: Performance Evaluation for Capillary Isoelectric Focusing.

Capillary electrophoresis (CE) combined with mass spectrometry (MS) is a powerful analytical technique that utilizes the resolving power of CE and the mass-detection capabilities of MS. In many cases, CE is coupled to MS via a sheath-flow interface (SFI). This interface has a simple design and can be easily constructed; however, it often suffers from issues such as MS signal suppression, interference of MS and CE electrical circuits, and the inability to set an optical point of detection close to the capillary end due to the specific design of the coupling union. In this paper, we describe a novel coupling of CE and MS based upon the open port interface (OPI). The OPI differs from classical sheath flow interfaces by operating at flow rates at least 1 order of magnitude higher. In addition to the flow rate difference, the OPI provides more efficient mixing of the capillary eluates with the transport fluid and thus minimizes MS signal suppression. In this work, we compared the performance of OPI and SFI in a series of capillary isoelectric focusing (cIEF) experiments with 5 pI markers, carbonic anhydrase II and NIST antibody. The evaluation criteria for the comparison of the OPI and SFI were analytical sensitivity, reproducibility, and pI marker linearity. Given the extent of sample dilution in the OPI, we also compared the peak resolution determined using an upstream UV detector to those determined by the downstream mass spectrometer. The results suggested that the OPI configuration reduced signal suppression, with no adverse effect on peak resolution. In addition, the OPI provided better decoupling of the CE and MS potentials as well as reduced signal dependence upon the sheath liquid composition. While these results are preliminary, they suggest that the OPI is a viable approach for CE-MS coupling.

3D printer platform and conductance feedback loop for automated imaging of uneven surfaces by liquid microjunction-surface sampling probe mass spectrometry.

RATIONALE: Molecular imaging of samples using mass spectrometric techniques, such as matrix-assisted laser desorption ionization or desorption electrospray ionization, requires the sample surface to be even/flat and sliced into thin sections (c. 10 µm). Furthermore, sample preparation steps can alter the analyte composition of the sample. The liquid microjunction-surface sampling probe (LMJ-SSP) is a robust sampling interface that enables surface profiling with minimal sample preparation. In conjunction with a conductance feedback system, the LMJ-SSP can be used to automatically sample uneven specimens. METHODS: A sampling stage was built with a modified 3D printer where the LMJ-SSP is attached to the printing head. This setup can scan across flat and even surfaces in a predefined pattern ("static sampling mode"). Uneven samples are automatically probed in "conductance sampling mode" where an electric potential is applied and measured at the probe. When the probe contacts the electrically grounded sample, the potential at the probe drops, which is used as a feedback signal to determine the optimal position of the probe for sampling each location. RESULTS: The applicability of the probe/sensing system was demonstrated by first examining the strawberry tissue using the "static sampling mode." Second, porcine tissue samples were profiled using the "conductance sampling mode." With minimal sample preparation, an area of 11 × 15 mm was profiled in less than 2 h. From the obtained results, adipose areas could be distinguished from non-adipose parts. The versatility of the approach was further demonstrated by directly sampling the bacteria colonies on agar and resected human kidney (intratumoral hemorrhage) specimens with thicknesses ranging from 1 to 4 mm. CONCLUSION: The LMJ-SSP in conjunction with a conductive feedback system is a powerful tool that allows for fast, reproducible, and automated assessment of uneven surfaces with minimal sample preparation. This setup could be used for perioperative assessment of tissue samples, food screening, and natural product discovery, among others.

Hyperspectral Visualization-Based Mass Spectrometry Imaging by LMJ-SSP: A Novel Strategy for Rapid Natural Product Profiling in Bacteria.

Mass spectrometry imaging (MSI) has been widely used to discover natural products (NPs) from underexplored microbiological sources. However, the technique is limited by incompatibility with complicated/uneven surface topography and labor-intensive sample preparation, as well as lengthy compound profiling procedures. Here, liquid micro-junction surface sampling probe (LMJ-SSP)-based MSI is used for rapid profiling of natural products from Gram-negative marine bacteria Pseudoalteromonas on nutrient agar media without any sample preparation. A conductance-based autosampling platform with 1 mm spatial resolution and an innovative multivariant analysis-driven method was used to create one hyperspectral image for the sampling area. NP discovery requires general spatial correlation between m/z and colony location but not highly precise spatial resolution. The hyperspectral image was used to annotate different m/z by straightforward color differences without the need to directly interrogate the spectra. To demonstrate the utility of our approach, the rapid analysis of Pseudoalteromonas rubra DSM6842, Pseudoalteromonas tunicata DSM14096, Pseudoalteromonas piscicida JCM20779, and Pseudoalteromonas elyakovii ATCC700519 cultures was directly performed on Agar. Various natural products, including prodiginine and tambjamine analogues, were quickly identified from the hyperspectral image, and the dynamic extracellular environment was shown with compound heatmaps. Hyperspectral visualization-based MSI is an efficient and sensitive strategy for direct and rapid natural product profiling from different Pseudoalteromonas strains.

Investigating electrochemical deposition of gold on commercial off-the-shelf 3-D printing materials towards developing sensing applications.

The COVID-19 pandemic highlighted the inaccessibility of quick and affordable clinical diagnostics. This led to increased interest in creating low-cost portable electrochemical (EC) devices for environmental monitoring and clinical diagnostics. One important perspective is to develop new fabrication methods for functional and low-cost electrode chips. Techniques, such as electron beam and photolithography, allow precise and high-resolution electrode fabrication; however, they are costly and can be time-consuming. More recently, fused deposition modeling three-dimensional (3-D) printing is being used as an alternative fabrication technique due to the low-cost of the printer and rapid prototyping capability. In this study, we explore enhancing the conductivity of 3-D printed working electrodes with EC gold deposition. Two commercial conductive filament brands were used and investigated to fabricate electrode chips. Furthermore, strategies to apply epoxy glue and conductive silver paint were investigated to control the electrode surface area and ensure good electrical connection. This device enables detection at drinking water concentration thresholds. The practical application of the fabricated electrodes is demonstrated by detecting Cu2+ using anodic stripping voltammetry.

Discontinuously Dewetting Solvent Arrays: Droplet Formation and Poly-Synchronous Surface Extraction for Mass Spectrometry Imaging Applications.

We describe a new liquid tissue stamping method called poly-synchronous surface extraction (PSSE) that utilizes an omniphobic substrate patterned with hydrophilic surface energy traps (SETs), which when wet with a solvent form a dense microdroplet array. When contacted with a tissue sample, each droplet locally extracts analytes from the tissue surface, which subsequentially can be analyzed by matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-IMS) or ambient ionization-MS techniques. Optimization of the patterned surface with six different solvents was carried out to increase the droplet density, height, and reproducibility of volume deposition. Once optimized, sister slices of a strawberry (Fragaria × ananassa) were spatially extracted using the PSSE technique and the chemical distribution of selected compounds was analyzed with both MALDI-IMS and a lower resolution but faster ambient liquid microjunction surface sampling probe (LMJ-SSP) approach. Heat maps for target analytes for the PSSE approach are compared to those produced using traditional MALDI-IMS analysis. The PSSE method aligned well with direct analysis and demonstrated the potential to increase the speed of ambient MS tissue imaging techniques by decreasing the number of steps required for sample preparation.

Rapid Mass Spectrometric Calibration and Standard Addition Using Hydrophobic/Hydrophilic Patterned Surfaces and Discontinuous Dewetting.

The rapid calibration chip (RCC) is a device that uses the fast and reproducible wetting behavior of hydrophilic/hydrophobic patterned surfaces to confine a series of differently sized droplets on a substrate to obtain a calibration curve. Multiple series of droplets can be formed within seconds by dipping an RCC into a calibration solution. No pipetting, sequential droplet deposition, or advanced equipment is required. The performance and reproducibility of RCCs were evaluated with an electrospray ionization triple-quadrupole mass spectrometer equipped with a liquid microjunction-surface sampling probe (LMJ-SSP) that allows for fast sampling of surfaces. Using circular hydrophilic areas with diameters ranging from 0.25 to 2.00 mm, liquid volumes of 4.6-70.6 nL could be deposited. Furthermore, the use of a second hydrophobic/hydrophilic patterned transfer chip can be used to add internal standard solutions to each calibration spot of the RCC, allowing to transfer a liquid volume of 22.5 nL.

Application of Intraoperative Mass Spectrometry and Data Analytics for Oncological Margin Detection, A Review.

OBJECTIVE: A common phase of early-stage oncological treatment is the surgical resection of cancerous tissue. The presence of cancer cells on the resection margin, referred to as positive margin, is correlated with the recurrence of cancer and may require re-operation, negatively impacting many facets of patient outcomes. There exists a significant gap in the surgeon's ability to intraoperatively delineate between tissues. Mass spectrometry methods have shown considerable promise as intraoperative tissue profiling tools that can assist with the complete resection of cancer. To do so, the vastness of the information collected through these modalities must be digested, relying on robust and efficient extraction of insights through data analysis pipelines. METHODS: We review clinical mass spectrometry literature and prioritize intraoperatively applied modalities. We also survey the data analysis methods employed in these studies. RESULTS: Our review outlines the advantages and shortcomings of mass spectrometry imaging and point-based tissue probing methods. For each modality, we identify statistical, linear transformation and machine learning techniques that demonstrate high performance in classifying cancerous tissues across several organ systems. A limited number of studies presented results captured intraoperatively. CONCLUSION: Through continued research of data centric techniques, like mass spectrometry, and the development of robust analysis approaches, intraoperative margin assessment is becoming feasible. SIGNIFICANCE: By establishing the relatively short history of mass spectrometry techniques applied to surgical studies, we hope to inform future applications and aid in the selection of suitable data analysis frameworks for the development of intraoperative margin detection technologies.

The liquid micro junction-surface sampling probe (LMJ-SSP); a versatile ambient mass spectrometry interface.

Ambient ionization methods have become important tools in mass spectrometry. The LMJ-SSP can significantly simplify/reduce lengthy sample preparation requirements associated with mass spectrometry analysis. Samples may be introduced through direct contact, insertion and droplet injection, enabling applications from drug discovery and surface analysis to tissue profiling and metabolic mapping. This review examines the underlying principles associated with the LMJ-SSP interface and highlights modifications of the original design that have extended its capability. We summarize different application areas that have exploited the method and describe potential future directions for the adaptable ambient ionization source.

Fabrication and characterization of laser-heated, multiplexed electrospray emitter.

We present a one-step fabrication method for a new multiplexed electrospray emitter with nine parallel micronozzles. The nozzles were formed by wet chemical etching of the end of a microstructured silica fiber containing nine 10 µm flow channels. By carefully adjusting the water flow through the channels while etching, we controlled the shape of the conical micronozzles and were able to obtain conditions under which the micronozzles, together with the flow channels, formed optical micro-axicon lenses. When 1064 nm light was guided through the flow channels and focused by the micro-axicon lenses into the Taylor cones, we were able to increase the desolvation of a model analyte and thereby increased the spray current produced by the emitter. This work paves the way towards a rapidly modulated mass-spectrometry source having a greatly enhanced throughput.

Ice recrystallization inhibition activity varies with ice-binding protein type and does not correlate with thermal hysteresis.

Ice-binding proteins (IBPs) inhibit the growth of ice through surface adsorption. In some freeze-resistant fishes and insects, circulating IBPs serve as antifreeze proteins to stop ice growth by lowering the freezing point. Plants are less able to avoid freezing and some use IBPs to minimize the damage caused in the frozen state by ice recrystallization, which is the growth of large ice grains at the expense of small ones. Here we have accurately and reproducibly measured the ice recrystallization inhibition (IRI) activity of over a dozen naturally occurring IBPs from fishes, insects, plants, and microorganisms using a modified 'splat' method on serial dilutions of IBPs whose concentrations were determined by amino acid analysis. The endpoint of IRI, which was scored as the lowest protein concentration at which no recrystallization was observed, varied for the different IBPs over two orders of magnitude from 1000 nM to 5 nM. Moreover, there was no apparent correlation between their IRI levels and reported antifreeze activities. IBPs from insects and fishes had similar IRI activity, even though the insect IBPs are typically 10x more active in freezing point depression. Plant IBPs had weak antifreeze activity but were more effective at IRI. Bacterial IBPs involved in ice adhesion showed both strong freezing point depression and IRI. Two trends did emerge, including that basal plane binding IBPs correlated with stronger IRI activity and larger IBPs had higher IRI activity.

Realizing new designs of multiplexed electrode chips by 3-D printed masks.

Creating small and portable analytical methods is a fast-growing field of research. Devices capable of performing bio-analytical detection are especially desirable with the onset of the global pandemic. Lab-on-a-chip (LOC) technologies, including rapid point-of-care (POC) devices such as glucose sensors, are attractive for applications in resource-poor settings. There are many challenges in creating such devices, from sensitive molecular designs to stable conditions for storing the sensor chips. In this study we have explored using three-dimensional (3D) printing to create shadow masks as a low-cost method to produce multiplexed electrodes by physical vapour deposition. Although the dimensional resolution of the electrodes produced by using 3D printed masks is inferior to those made through photolithography-based techniques, their dimensions can be readily tailored ranging from 1 mm to 3 mm. Multiple mask materials were tested, such as polylactic acid and polyethylene terephthalate glycol, with acrylonitrile butadiene styrene shown to be the best. Simple strategies in making chip holders by 3D printing and controlling working electrode surface area with epoxy glue were also investigated. The prepared chips were tested by performing surface chemistry with thiol-containing molecules and monitoring the signals electrochemically.

Detection of Opioids on Mail/Packages Using Open Port Interface Mass Spectrometry (OPI-MS).

Opioids (and their more potent synthetic analogues) are used therapeutically as effective pain killers; however, recreational use and consequent overdoses are implicated in the deaths of thousands of people across the world annually. Trafficking of opioids and other illegal drugs through international mail has become a significant challenge for law enforcement personnel. Hundreds of millions of letters are sorted by the U.S. and Canadian postal services every day. Chemical analysis of this immense volume of mail requires a very fast sampling/detection method. This work explores the use of real-time mass spectrometry analysis with the recently developed Open Port Interface (OPI) for acoustically dispensed nanoliter volume sample droplets, a type of liquid microjunction surface sampling probe, for rapid and easy non-intrusive detection of fentanyl, heroin, and oxycodone. The OPI coupled to mass spectrometry is a novel sample introduction method that allows the rapid analysis of sample surfaces without preparation or modification. Opioids on different packaging materials (e.g., paper, bubble wrap, Ziploc bags) were rapidly (<10 s) interrogated by the OPI, and the sensitivities of the method compared. Furthermore, an opioid surrogate (caffeine) could be facilely detected on envelopes after processing through postal services.

Open sessile droplet viscometer with low sample consumption.

This paper reports a portable viscometer that requires less than 10 µL of sample for a measurement. Using a two-droplet Laplace-induced pumping system on an open microfluidic substrate, the device measures the viscosity of a liquid by determining the time required for one droplet to completely pump into a second droplet. The pumping behaviour follows the Hagen-Poiseuille and Laplace relations where the flow rate, Q, is proportional to the liquid's kinematic viscosity, µ. The progress of pumping is measured by tracking the change in curvature of one of the droplets using a laser that is positioned perpendicular to the microfluidic chip and directed at the "tail" of the shrinking droplet. The angle of incidence and degree of refraction changes depending on the size of the droplet, which is tracked by a linear diode array placed beneath the microfluidic chip. Droplet reservoirs and connecting channels were defined by precise patterning of a glass substrate coated with a commercially available omniphobic coating (Ultra Ever Dry®) using laser micromachining. A 500 µm wide and 20 mm long channel with circular reservoirs (d = 1.5 mm) enabled the measurement of dynamic viscosities in the range of η = 1.0-2.87 mPa s. The materials cost for the entire viscometer (fluidics and electronics, etc.) is <15 USD.

Hydrophobic/hydrophilic patterned surfaces for directed evaporative preconcentration.

The deposition of micro- and nanolitre volumes is crucial in sessile droplet microfluidic systems. Several techniques exist for the fabrication of surfaces with patterned wettabilities; however, many of these fabrication techniques are time-consuming and complex. Here, we present a device that allows for deposition of multiple droplets within seconds followed by directed evaporative preconcentration. Hydrophobic-coated glass substrates are fashioned with hydrophilic surface energy traps (SETs) using picosecond laser micromachining. SETs can capture nanolitre volumed droplets of both aqueous and organic liquids through discontinuous dewetting. Modification of the machined hydrophilic shape yields a passive mechanism that preconcentrates analyte through evaporation. Studies and optimizations of SET parameters/dimensions (laser power, laser passes, ring/patch diameter) and their effect on patch wettability and degree of preconcentration are presented. As a demonstration, the optimized platform was used to improve the colourimetric detection of cadmium-containing aqueous samples. The optimized SET design demonstrated an 18-fold increase in colourimetric sensitivity compared to conventional milled SETs, suggesting the design would be well-suited for trace analysis. The evaporative preconcentration was also applied to MALDI-IMS analysis of peptides where it resulted in improved uniformity of deposited analyte and decreased analysis times. The rapid droplet deposition and directed evaporative approach can be tailored to provide different concentration factors and is compatible with a wide variety of detection schemes.

Light activated synthesis of the atomically precise fluorescent silver cluster Ag18(Capt)14.

Metal clusters of gold and silver with highly tunable optical and electronic properties are attractive candidates for next generation medical imaging and therapy. Of these two most commonly studied metals, silver clusters often exhibit superior optical properties (i.e. stronger absorbance and higher emission quantum yield). The atomically precise synthesis of these clusters is essential before their use in biological applications can be realized. However, most cluster synthetic routes result in complex mixtures, where isolation and/or characterization can become incredibly challenging. Using photochemistry, we demonstrate a synthetic route for silver thiolate clusters resulting in the isolation of a pure eighteen-atom silver cluster capped by fourteen captopril ligands, Ag18(Capt)14. The facile control over the reduction of Ag(i) salt that this photochemical route affords can be readily applied as a general synthesis for isolating other new, atomically precise clusters.

Facile actuation of aqueous droplets on a superhydrophobic surface using magnetotactic bacteria for digital microfluidic applications.

Magnetic actuation provides a low-cost, simple method for droplet manipulation on a digital microfluidic platform. The impetus to move the droplets on a low friction surface can come from internal superparamagnetic particles or paramagnetic salts. Recently, the use of microbes for bio-actuation has been established, where the thrust produced by the microbes can be exploited to exert the force required for droplet movement. This study presents biologically-driven magnetic actuation of droplets on a superhydrophobic surface using magnetotactic bacteria (MTB). MTB-droplets were impelled along various trajectories such as rectangular and figure-of-eight-shaped paths. Droplets were reproducibly actuated with speeds up of to 30 mm s-1. We demonstrated the ability to sequentially merge and mix multiple droplets by merging a 10 µL MTB droplet with two 4 µL colored droplets. The reorientation of MTB in the droplet enhanced mixing rate of the merged fluids by ∼40% compared with the control experiment where no actuation was used. Biologically-driven magnetic actuation was compared with actuation by superparamagnetic particles and paramagnetic salts, in terms of controllability and speed. MTB droplet was moved with the same average speed as other two methods and showed higher response time as the magnet acceleration increased. Lastly, MTB were used to perform a phosphatase assay using endogenous enzyme. The relative absorbance at 405 nm, indicating the production of the yellow product, increased over time and levels off after 75 min.

Facile Actuation of Organic and Aqueous Droplets on Slippery Liquid-Infused Porous Surfaces for the Application of On-Chip Polymer Synthesis and Liquid-Liquid Extraction.

Digital microfluidics employs water-repellant surfaces to exquisitely manipulate droplets of water for chemical analysis. However, the actuation and manipulation of organic droplets is still relatively unexplored as it is significantly more difficult to synthesize organic-repellent surfaces compared to water-repellent surfaces. Here, we present the fabrication of slippery liquid-infused porous surfaces (SLIPS) based on a porous polymer monolithic approach. The synthesized SLIPS were able to repel organic liquids such as hexane and methanol with a contact angle of 42.1 ± 0.4° and 69.0 ± 1.8°, respectively, as well as water with a contact angle of 115.8 ± 0.8°. More importantly for digital microfluidic applications, the sliding angle of liquids tested was between 4° and 6°. As a result, droplets containing magnetically susceptible material could be facilely manipulated on the SLIPS surface. A systematic actuation study was carried out to explore how actuation parameters including speed, paramagnetic particle (PMP) concentrations, and droplet volume impacted the outcomes (droplet actuation, disengagement, and PMP extraction). Two different applications were used to demonstrate the utility of actuating organic droplets on SLIPS surfaces including on-chip liquid-liquid extractions of natural products (NPs) from marine bacteria and droplet-based polymer synthesis with different polymerization conditions. Both applications employ an aqueous droplet and organic droplet interface at which either phase transfer or a chemical reaction is carried out. Two NPs (prodigiosin from Pseudoalteromonas rubra and violacein from Pseudoalteromonas luteoviolacea) were extracted, from aqueous droplets containing the bacteria, into butanol droplets and characterized with matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS). Nylon 6,6 was synthesized on-chip via magnetic actuation of organic droplets containing adipoyl chloride and hexamethylamine. Relative intensities of the characteristic polymer masses suggest that droplet-based microfluidic synthesis on slips can be used to probe reaction conditions. The compatibility of SLIPS with both aqueous and organic solutions opens up a wider number of droplet-based sample preparation protocols and chemical transformations.

Reliable identification of prostate cancer using mass spectrometry metabolomic imaging in needle core biopsies.

Metabolomic profiling can aid in understanding crucial biological processes in cancer development and progression and can also yield diagnostic biomarkers. Desorption electrospray ionization coupled to mass spectrometry imaging (DESI-MSI) has been proposed as a potential adjunct to diagnostic surgical pathology, particularly for prostate cancer. However, due to low resolution sampling, small numbers of mass spectra, and little validation, published studies have yet to test whether this method is sufficiently robust to merit clinical translation. We used over 900 spatially resolved DESI-MSI spectra to establish an accurate, high-resolution metabolic profile of prostate cancer. We identified 25 differentially abundant metabolites, with cancer tissue showing increased fatty acids (FAs) and phospholipids, along with utilization of the Krebs cycle, and benign tissue showing increased levels of lyso-phosphatidylethanolamine (PE). Additionally, we identified, for the first time, two lyso-PEs with abundance that decreased with cancer grade and two phosphatidylcholines (PChs) with increased abundance with increasing cancer grade. Importantly, we developed and internally validated a multivariate metabolomic classifier for prostate cancer using 534 spatial regions of interest (ROIs) in the training cohort and 430 ROIs in the test cohort. With excellent statistical power, the training cohort achieved a balanced accuracy of 97% and validation on testing data set demonstrated 85% balanced accuracy. Given the validated accuracy of this classifier and the correlation of differentially abundant metabolites with established patterns of prostate cancer cell metabolism, we conclude that DESI-MSI is an effective tool for characterizing prostate cancer metabolism with the potential for clinical translation.