Bioanalytical Tools Research Articles

New incorporation of the S9 metabolizing system into methods for detecting acetylcholinesterase inhibition

The activity of individual biotransformation products cannot be measured in multicomponent mixtures by the current status-quo assays. A prior separation and tedious isolation of compounds is required, and often in addition, a concentration step into a solvent suitable for the cell-/enzyme-based assay. Hence, the metabolizing S9 system, mimicking the complex biotransformation reactions in the liver, was aimed to be integrated into two orthogonal methods for analysis of the acetylcholinesterase (AChE) inhibition. For the microtiter plate assay method, the evaluation of the generated fluorescence signal was impaired by the incorporated S9 system. In contrast, the metabolic activator (S9 mixture) was successfully incorporated into the high-performance thin-layer chromatography (HPTLC) method. As proof of principle, four reference AChE inhibitors were studied in complex samples with and without metabolic activation. In addition to the neurotoxic carbamate eserine and the organophosphate insecticides chlorpyrifos, quinalphos and parathion, the tris(nonylphenyl) phosphite and nonylphenol, both originating from food contact materials, were tested in isolation but also in food packaging migrate and extract. A method comparison and benchmarking pointed to multifold advantages of using this newly developed bioanalytical tool for assessment of individual neurotoxins in complex samples. The sensitive HPTLC-S9-AChE assay allowed the detection of neurotoxic chemicals with and without metabolic activation, at levels consistent with the threshold of toxicological concern of organophosphates and carbamates. This new on surface metabolism system can be applied to other toxicities and samples.

Microbial electrochemical biosensor for rapid detection of naphthenic acid in aqueous solution

Abstract In processed water generated from oil sands operations, naphthenic acids (NAs) are identified as one of the primary sources of acute toxicity and were assessed for their endocrine-disrupting potentials. Hence, the monitoring of NAs in aqueous environmental matrices is crucial for oil sands and crude oil production industries. Commonly used analytical methods for NAs include Fourier transform infrared spectroscopy, gas chromatography–mass spectrometry, and high-performance liquid chromatography. However, these methods are time-consuming, expensive, and samples must be sent to an analytical lab for analysis. In this study, we first demonstrated microbial electrochemical cell (MXC)-based biosensor for the rapid detection of a model NA compound (cyclohexane carboxylic acid) in water samples. Under the continuous closed-circuit operation of MXC biosensor, electrical responses (peak currents) were fairly proportional (R2 = 0.64) to the model NA concentrations ranging from 50 to 250 mg COD/L. However, the charging-discharging operation of biosensor significantly increased the peak currents by 90–124 folds higher than that observed for continuous closed-circuit operation. Besides, a linear relationship between model NA concentrations and the peak currents (R2 = 0.96) was observed. Furthermore, it has been revealed that the biosensor would be sensitive to salinity levels and temperature changes; however, once calibrated it can be used for measurement of model NA concentrations. Overall, the results of this study demonstrated that with further development, MXC biosensor could be used as a simple bioanalytical tool for monitoring NA concentrations in oil sands process-affected water.

BIOANALYSIS AND BIOTRANSFORMATION OF OLIGONUCLEOTIDE THERAPEUTICS BY LIQUID CHROMATOGRAPHY-MASS SPECTROMETRY.

Since 2016, eight new oligonucleotide therapies have been approved which has led to increased interest in oligonucleotide analysis. There is a particular need for powerful bioanalytical tools to study the metabolism and biotransformation of these molecules. This review provides the background on the biological basis of these molecules as currently used in therapies. The article also reviews the current state of analytical methodology including state of the art sample preparation techniques, liquid chromatography-mass spectrometry methods, and the current limits of detection/quantitation. Finally, the article summarizes the challenges in oligonucleotide bioanalysis and provides future perspectives for this emerging field. © 2020 John Wiley & Sons Ltd.

Orthogonal Hyphenation of Planar and Liquid Chromatography for Mass Spectrometry of Biomarkers out of the Bioassay Matrix (NP-HPTLC-UV/vis/FLD-Bioassay-RP/IEX-HPLC-UV/vis-ESI-MS).

Bioprofiling on the planar chromatogram with in situ biological/enzymatic assays is a powerful bioanalytical screening tool for the nontargeted detection of known and especially unknown/unidentified bioactive compounds directly in multicomponent mixtures (e.g., foods, spices, and botanicals). However, together with the bioactive zone, the adsorbed bioassay medium is eluted into the mass spectrometer (MS) and interfering with evaluation. Another sample track without bioassay has thus been handled in parallel. Hence, for a direct zone elution from the bioautogram, different setups were investigated to reduce the impact of the bioassay medium load. A biocompatible filter, orthogonal reversed-phase/cation-exchange columns (RP/IEX-HPLC), UV/vis detector, and a Rheodyne valve were installed between the zone eluting interface (after normal-phase high-performance thin-layer chromatography-multi-imaging-bioassay, NP-HPTLC-UV/vis/FLD-bioassay) and the MS. For the negative electrospray ionization mode (ESI-), an RP-18e-HPLC column and valve switch were exploited. After gradient optimization, the RP-column retarded the eluted polar compounds and split-off the salts of the bioassay medium in the first minutes. This reduced the bioassay load and separated analyte signals thereof. However, most bioassay medium mass signals were predominantly detectable in ESI+-MS. Here, the reduction of bioassay matrix signals was achieved by integrating a mixed-mode RP/IEX column. Finally, two different superhyphenations were successfully proven: NP-HPLC-UV/vis/FLD-bioassay-RP-HPLC-UV/vis-ESI--MS with a valve switch and NP-HPLC-UV/vis/FLD-bioassay-RP/IEX-HPLC-UV/vis-ESI±-MS with or without it. Although the original bioprofiling (NP-HPTLC-UV/vis/FLD-bioassay) was prolonged from 3 to 13 min per sample, such superhyphenations covering chemistry/biology/mass spectrometry are considered as an efficient nontarget bioanalytical tool for fast evaluation of complex samples.

Non-targeted detection and differentiation of agonists versus antagonists, directly in bioprofiles of everyday products

Xenoestrogens exert antiandrogenic effects on the human androgen receptor. In the analytical field, such antagonists block the detection of testosterone and falsify results obtained by sum parameter assays. Currently, such agonistic versus antagonistic effects are not differentiated in complex mixtures. Oppositely acting hormonal effects present in products of everyday use can only be differentiated after tedious fractionation and isolation of the individual compounds along with subjection of each fraction/compound to the status quo bioassay testing. However, such long-lasting procedures are not suited for routine. Hence, we developed a fast bioanalytical tool that figures out agonists versus antagonists directly in complex mixtures. Exemplarily, 8 cosmetics and 15 thermal papers were analyzed. The determined antagonistic potentials of active compounds found were comparable to the ones of known antagonists (in reference shown for bisphenol A, 4-n-nonylphenol and four parabens). Relevant biological/chromatographic parameters such as cell viability, culture conditions, dose response curves, limits of biological detection/quantification and working range (shown for testosterone, dihydrotestosterone, nandrolone and trenbolone) were investigated to obtain the best sensitivity of the biological detection. The developed and validated method was newly termed reversed phase high-performance thin-layer chromatography planar yeast ant-/agonistic androgen screen (RP-HPTLC-pYAAS bioassay). Results were also compared with the RP-HPTLC-Aliivibrio fischeri bioassay (applied on RP plates for the first time). As proof-of-concept, the transfer to another bioassay (RP-HPTLC-pYES) was successfully demonstrated, analogously termed RP-HPTLC-pYAES bioassay detecting anti-/estrogens (exemplarily shown for evaluation of 4 pharmaceuticals used in breast cancer treatment). The new imaging concept provides (1) detection and differentiation of individual agonistic versus antagonistic effects in the bioprofiles, (2) bioanalytical quantification of their activity potential by scanning densitometry and (3) characterization of unknown bioactive compound zones by hyphenation to high-resolution mass spectrometry. Depending on the hormonal bioassay, 15 samples were analyzed in parallel within 5 h or 6 h (calculated as 20 or 24 min per sample). For the first time, piezoelectric spraying of the yeast cells was successfully demonstrated for the planar yeast–based bioassays.

Cannabinomics: Application of Metabolomics in Cannabis (Cannabis sativa L.) Research and Development.

Cannabis (Cannabis sativa L.) is a complex, polymorphic plant species, which produces a vast array of bioactive metabolites, the two major chemical groups being cannabinoids and terpenoids. Nonetheless, the psychoactive cannabinoid tetrahydrocannabinol (Δ9-THC) and the non-psychoactive cannabidiol (CBD), are the two major cannabinoids that have monopolized the research interest. Currently, more than 600 Cannabis varieties are commercially available, providing access to a multitude of potent extracts with complex compositions, whose genetics are largely inconclusive. Recently introduced legislation on Cannabis cultivation in many countries represents a great opportunity, but at the same time, a great challenge for Cannabis research and development (R&D) toward applications in the pharmaceutical, food, cosmetics, and agrochemical industries. Based on its versatility and unique capabilities in the deconvolution of the metabolite composition of complex matrices, metabolomics represents an ideal bioanalytical tool that could greatly assist and accelerate Cannabis R&D. Among others, Cannabis metabolomics or cannabinomics can be applied in the taxonomy of Cannabis varieties in chemovars, the research on the discovery and assessment of new Cannabis-based sources of bioactivity in medicine, the development of new food products, and the optimization of its cultivation, aiming for improvements in yield and potency. Although Cannabis research is still in its infancy, it is highly foreseen that the employment of advanced metabolomics will provide insights that could assist the sector to face the aforementioned challenges. Within this context, here, the current state-of-the-art and conceptual aspects of cannabinomics are presented.

Plasmonic nanoparticles assemblies templated by helical bacteria and resulting optical activity.

Plasmonic nanoparticles (NPs) adsorbing onto helical bacteria can lead to formation of NP helicoids with micron scale pitch. Associated chiroptical effects can be utilized as bioanalytical tool for bacterial detection and better understanding of the spectral behavior of helical self-assembled structures with different scales. Here, we report that enantiomerically pure helices with micron scale of chirality can be assembled on Campylobacter jejuni, a helical bacterium known for severe stomach infections. These organisms have right-handed helical shapes with a pitch of 1-2 microns and can serve as versatile templates for a variety of NPs. The bacteria itself shows no observable rotatory activity in the visible, red, and near-IR ranges of electromagnetic spectrum. The bacterial dispersion acquires chiroptical activity at 500-750 nm upon plasmonic functionalization with Au NPs. Finite-difference time-domain simulations confirmed the attribution of the chiroptical activity to the helical assembly of gold nanoparticles. The position of the circular dichroism peaks observed for these chiral structures overlaps with those obtained before for Au NPs and their constructs with molecular and nanoscale chirality. This work provides an experimental and computational pathway to utilize chiroplasmonic particles assembled on bacteria for bioanalytical purposes.

Comprehensive Bioanalysis of Ultrahigh Molecular Weight, Highly Disperse Poly(ethylene oxide) in Rat via Microsolid Phase Extraction and RPLC-Q-Q-TOF Coupled with the MSALL Technique.

Ultrahigh molecular weight (UHMW) poly(ethylene oxide) (PEO) is a synthetic hydrophilic polymer with wide dispersity which shows considerable promise as a hemostatic agent in the treatment of gastrointestinal bleeding. Currently there is no analytical method for the determination of highly disperse UHMW PEO in biological samples that would allow its characterization in vivo and support its clinical development. Although liquid chromatography-tandem mass spectrometry (LC-MS/MS) is a powerful bioanalytical tool, it faces major challenges when applied to UHMW PEO. In this work, we report a novel bioanalytical method for the determination of UHMW PEO involving microsolid phase extraction (μ-SPE), chromatography on a PLRP-S 1000 Å reversed phase column and detection by positive ion Q-Q-TOF MS using the MSALL technique. In this mode, dissociation of all precursor ions in Q2 generated a series of product ions at m/z 89.0715, 133.0854, 177.1047, and 221.1475 of which the product ion at m/z 133.0854 was common to all precursor ions and enabled quantitation of all polymers in UHMW PEO. The method was successfully applied to the determination of UHMW PEO polymers in rat plasma, urine, and feces after oral administration of 1700 kDa PEO. The results show that UHMW PEO is not absorbed into the blood and is largely eliminated unchanged in feces over 48 h. We maintain the method is sufficiently robust to be used in routine bioanalysis of polymers with UHMW and wide dispersity.

Addressing the Analytical Challenges for the Detection of Ciguatoxins Using an Electrochemical Biosensor.

The importance of ciguatoxins (CTXs) in seafood safety and their emerging occurrence in locations far away from tropical areas highlight the need for simple and low-cost methods for the sensitive and rapid detection of these potent marine toxins to protect seafood consumers. Herein, an electrochemical immunosensor for the detection of CTXs is presented. A sandwich configuration is proposed, using magnetic beads (MBs) as immobilization supports for two capture antibodies, with their combination facilitating the detection of CTX1B, CTX3C, 54-deoxyCTX1B, and 51-hydroxyCTX3C. PolyHRP-streptavidin is used for the detection of the biotinylated detector antibody. Experimental conditions are first optimized using colorimetry, and these conditions are subsequently used for electrochemical detection on electrode arrays. Limits of detection at the pg/mL level are achieved for CTX1B and 51-hydroxyCTX3C. The applicability of the immunosensor to the analysis of fish samples is demonstrated, attaining detection of CTX1B at contents as low as 0.01 μg/kg and providing results in correlation with those obtained using mouse bioassay (MBA) and cell-based assay (CBA), and confirmed by liquid chromatography coupled to high-resolution mass spectrometry (LC-ESI-HRMS). This user-friendly bioanalytical tool for the rapid detection of CTXs can be used to mitigate ciguatera risk and contribute to the protection of consumer health.

Classification of analytics, sensorics, and bioanalytics with polyelectrolyte multilayer capsules

Polyelectrolyte multilayer (PEM) capsules, constructed by LbL (layer-by-layer)-adsorbing polymers on sacrificial templates, have become important carriers due to multifunctionality of materials adsorbed on their surface or encapsulated into their interior. They have been also been used broadly used as analytical tools. Chronologically and traditionally, chemical analytics has been developed first, which has long been synonymous with all analytics. But it is not the only development. To the best of our knowledge, a summary of all advances including their classification is not available to date. Here, we classify analytics, sensorics, and biosensorics functionalities implemented with polyelectrolyte multilayer capsules and coated particles according to the respective stimuli and application areas. In this classification, three distinct categories are identified: (I) chemical analytics (pH; K+, Na+, and Pb2+ ion; oxygen; and hydrogen peroxide sensors and chemical sensing with surface-enhanced Raman scattering (SERS)); (II) physical sensorics (temperature, mechanical properties and forces, and osmotic pressure); and (III) biosensorics and bioanalytics (fluorescence, glucose, urea, and protease biosensing and theranostics). In addition to thisclassification, we discuss also principles of detection using theabove-mentioned stimuli.These application areas are expected to grow further, but the classification provided here should help (a) to realize the wealth of already available analytical and bioanalytical tools developed with capsules using inputs of chemical, physical, and biological stimuli and (b) to position future developments in their respective fields according to employed stimuli and application areas. Graphical abstract.

Evaluation of reverse osmosis drinking water treatment of riverbank filtrate using bioanalytical tools and non-target screening

Organic micropollutants that occurred in a natural drinking water source induced effects that were not detectable after reverse osmosis. Bioactive compounds were characterised by non-target screening of LC-HRMS data using open cheminformatics approaches.

Exploring Cancer Metabolism: Applications of Metabolomics and Metabolic Phenotyping in Cancer Research and Diagnostics.

Altered metabolism is one of the key hallmarks of cancer. The development of sensitive, reproducible and robust bioanalytical tools such as Nuclear Magnetic Resonance Spectroscopy and Mass Spectrometry techniques offers numerous opportunities for cancer metabolism research, and provides additional and exciting avenues in cancer diagnosis, prognosis and for the development of more effective and personalized treatments. In this chapter, we introduce the current state of the art of metabolomics and metabolic phenotyping approachesin cancer research and clinical diagnostics.

Peptide and protein modified metal clusters for cancer diagnostics.

The biomedical features of metal clusters have been explored in tumor diagnostic applications in recent years. Peptide or protein protected metal clusters with low toxicity, ultra-small size and good biocompatibility are ideal bioanalytical tools, and exhibit better cancer diagnostic properties that have been attractive to oncologists. This perspective provides a rigorous but succinct overview of cancer diagnosis as a working concept for metal clusters by reporting the latest significant advances in the applications of metal clusters in tumor-related bioanalysis and diagnosis. The materials design principles, bioanalytical mechanisms and biomedical applications of metal clusters are described, and then the potential challenges and prospects of metal clusters in cancer diagnosis are discussed. A perspective addressing the role of metal clusters in this field is required to understand their effects and functions, as well as for the scientific community to further advance the development of metal clusters for broader diagnostic applications.

Validation and implementation of an ultrasensitive liquid chromatographic-tandem mass spectrometric (LC-MS/MS) assay for dapivirine quantitation in breast milk

BackgroundThe non-nucleoside reverse transcriptase inhibitor dapivirine has been evaluated as a topical microbicidal agent to prevent HIV-1 acquisition. Several clinical trials have evaluated the pharmacokinetics of dapivirine when administered as a 25-mg intravaginal ring. Recent studies have focused on the distribution of dapivirine into breast milk. Drug distribution during lactation and breastfeeding can have implications in terms of infant drug exposure. Thus, sensitive bioanalytical tools are required to characterize the pharmacokinetics of dapivirine in breast milk. MethodsWhole breast milk was spiked with dapivirine and internal standard. Lipid content was disrupted via pre-treatment with n-hexane, and supernatants were subjected to solid phase extraction. Extracted materials were subjected to liquid chromatographic-tandem mass spectrometric (LC-MS/MS) analysis. Separation occurred using a Waters BEH C8, 50 × 2.1 mm UPLC column with a 1.7 µm particle size and dapivirine was detected on an API 5000 mass analyzer. Methods were validated in accordance with FDA Bioanalytical Method Validation recommendations. ResultsThe analytical method was optimized for dapivirine extraction from breast milk. The analytical measuring range of the assay was 10–1000 pg/mL. Calibration curves were generated via weighted linear regression of standards. Intra- and inter-assay precision and accuracy studies demonstrated %CVs ≤ 14.6% and %DEVs ≤ ±12.7%. Stability and matrix effects studies were also conducted and deemed acceptable. The method was applied to a previously reported phase 1 clinical trial and demonstrated appropriate performance in the quantitation of dapivirine in breast milk samples from lactating women. ConclusionsAn ultrasensitive LC-MS/MS assay has been developed and validated for the quantitation of dapivirine in breast milk. The described method meets validation acceptance criteria and has been applied to a phase 1 clinical trial.

Solid-Supported Proteins in the Liquid Chromatography Domain to Probe Ligand-Target Interactions.

Ligand-target interactions play a central role in drug discovery processes because these interactions are crucial in biological systems. Small molecules-proteins interactions can regulate and modulate protein function and activity through conformational changes. Therefore, bioanalytical tools to screen new ligands have focused mainly on probing ligand-target interactions. These interactions have been evaluated by using solid-supported proteins, which provide advantages like increased protein stability and easier protein extraction from the reaction medium, which enables protein reuse. In some specific approaches, precisely in the ligand fishing assay, the bioanalytical method allows the ligands to be directly isolated from complex mixtures, including combinatorial libraries and natural products extracts without prior purification or fractionation steps. Most of these screening assays are based on liquid chromatography separation, and the binding events can be monitored through on-line or off-line methods. In the on-line approaches, solid supports containing the immobilized biological target are used as chromatographic columns most of the time. Several terms have been used to refer to such approaches, such as weak affinity chromatography, high-performance affinity chromatography, on-flow activity assays, and high-performance liquid affinity chromatography. On the other hand, in the off-line approaches, the binding event occurs outside the liquid chromatography system and may encompass affinity and activity-based assays in which the biological target is immobilized on magnetic particles or monolithic silica, among others. After the incubation step, the supernatant or the eluate from the binding assay is analyzed by liquid chromatography coupled to various detectors. Regardless of the selected bioanalytical approach, the use of solid supported proteins has significantly contributed to the development of automated and reliable screening methods that enable ligands to be isolated and characterized in complex matrixes without purification, thereby reducing costs and avoiding time-laborious steps. This review provides a critical overview of recently developed assays.

High-Throughput Bioluminescence Imaging and Reporter Gene Assay with 3D Spheroids from Human Cell Lines.

3D cell culture models represent an attractive approach to decode intracellular and intercellular signaling, providing biologically relevant information and predictive data. Bioluminescent reporter gene assays and bioluminescence imaging in 3D cell models are very promising bioanalytical tools for several applications.Here we report a very straightforward method for bioluminescence imaging and bioluminescent reporter gene assays in 3D cell-culture models. Both the assays can be easily implemented in laboratories equipped with basic cell culture facilities and instrumentation for bioluminescence detection, that is, low-light detectors connected to inverted microscopes and luminometers, without the need for additional equipment.

Integration of Flow Cytometry and Single Cell Sequencing

Integrating cytometric analysis of cells, mitochondria, and other polynucleotide-containing biological particles with high-throughput single particle sequencing would provide an ultimate bioanalytical tool, simultaneously assessing phenotype, functionality, genome, and transcriptome of each particle in a large population. Here, we describe how such integration could be performed by adapting existing, well-established technologies.

Distinction and valorization of 30 root extracts of five goldenrod (Solidago) species

A high-performance thin-layer chromatography (HPTLC) method was developed for rapid and easy-to-perform discrimination between five goldenrod species present in Europe: the native Solidago virgaurea and the four invasive aliens, S. canadensis, S. gigantea, S. rugosa and S. graminifolia. The chemotaxonomic distinction was based on the chemical profile of their root extracts, confirmed by principal component analysis. This allowed the distinction of the goldenrods in wintertime, when classical morphological methods are not applicable. Their enzyme inhibitory profiles were determined by HPTLC combined with α-glucosidase, β-glucosidase, α-amylase, acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) assays. Two compounds of S. canadensis showed the most intense enzyme inhibition in all assays, having also antibacterial activity against Bacillus subtilis, Xanthomonas euvesicatoria and Aliivibrio fischeri strains. HPTLC-high-resolution mass spectrometry (HRMS), bioassay-guided isolation, NMR spectroscopy and literature data led to the characterization and identification of the labdane diterpenes solidagenone and presolidagenone as the active S. canadensis root components. The previously identified polyacetylenes (2Z,8Z and 2E,8Z matricaria esters) of S. virgaurea, also inhibited all enzymes. Except for the known anti-AChE effect of the 2Z,8Z-matricaria ester, this is the first report on the α-glucosidase, β-glucosidase, α-amylase, AChE and BChE inhibitory activity of these potent compounds. The anti-hyperglycemic effects of the S. canadensis labdanoids were also observed for the first time. Combined with effect-directed assays and HRMS, hyphenated HPTLC allowed an effect-directed high-throughput screening and a fast characterization of multipotent compounds. The investigation of botanicals by fast, hyphenated, bioanalytical tools substantially increased the information gain with regard to active principles (bioprofiling) and efficiently pointed to potent candidates for drug development.

Pneumatic unidirectional cell stretching device for mechanobiological studies of cardiomyocytes

In this paper, we present a transparent mechanical stimulation device capable of uniaxial stimulation, which is compatible with standard bioanalytical methods used in cellular mechanobiology. We validate the functionality of the uniaxial stimulation system using human-induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs). The pneumatically controlled device is fabricated from polydimethylsiloxane (PDMS) and provides uniaxial strain and superior optical performance compatible with standard inverted microscopy techniques used for bioanalytics (e.g., fluorescence microscopy and calcium imaging). Therefore, it allows for a continuous investigation of the cell state during stretching experiments. The paper introduces design and fabrication of the device, characterizes the mechanical performance of the device and demonstrates the compatibility with standard bioanalytical analysis tools. Imaging modalities, such as high-resolution live cell phase contrast imaging and video recordings, fluorescent imaging and calcium imaging are possible to perform in the device. Utilizing the different imaging modalities and proposed stretching device, we demonstrate the capability of the device for extensive further studies of hiPSC-CMs. We also demonstrate that sarcomere structures of hiPSC-CMs organize and orient perpendicular to uniaxial strain axis and thus express more maturated nature of cardiomyocytes.

Advances in surface‐enhanced Raman spectroscopy for cancer diagnosis and staging

AbstractSurface‐enhanced Raman spectroscopy (SERS) is rapidly emerging as a bioanalytical tool for cancer diagnosis and therapy. The SERS‐based molecular diagnostics have progressed from proof‐of‐concept studies towards analysis in animal models as well as for in vitro clinical diagnostics in the last decade. Recently, SERS has also been implemented in screening, diagnosis, and staging of clinical cancer samples. Moreover, in vivo SERS imaging has been implemented for mapping the extent of tumor growth and metastasis; SERS nanoparticles have also enabled image‐guided therapies strongly indicating SERS technology can significantly complement the practice of oncology. Despite the progress, widespread clinical translation of SERS nanoparticles is still challenging. Current SERS strategies in diagnostic oncology require further development and standardization to progress from bench‐top to point‐of‐care applications. The present review critically analyzes the current state of the art about various strategies for SERS‐based cancer detection and staging from cellular metabolites, exosomes, circulating tumor cells, extracellular fluids, and cancer cells.