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Spark® Cyto

Plate reader for 2D and 3D cell experiments with live cell imaging and real-time cytometry

Spark Cyto is a multi-mode plate reader with fluorescence imaging and cytometry capabilities, unlocking new possibilities for your cell-based research. By combining live cell imaging with industry-leading detection technologies, you now have the ability to unite qualitative and quantitative information into unique multi-parameter data sets.

Spark Cyto works in real -time, using parallel data acquisition and analysis- to deliver meaningful insights faster than before.

Revolutionize your cell experiments to never miss a critical biological event.

Tab 01 / Overview

Spark Cyto 3D AI

More insights delivered in real-time, uncover new approaches with Spark Cyto

More cells analyzed

Spark Cyto uniquely brings together top-of-the-range camera components with proprietary patent-pending technology to ensure that you can truly investigate your entire cell population. It gives you the ability to record the whole well area of a 96- or 384-well microplate with just one image, without tiling or distortion. This means that you never miss a cell when investigating the total cell population in a microplate well. Spark Cyto includes three magnification levels combined with four channels for fluorescence and bright field imaging, enabling high quality cell analysis for a wide variety of applications.

Learn more about the cell imaging module

Launching Spark Cyto at SLAS Europe 2019 - Lynda O'Leary

More parameters measured

Spark Cyto is available in configurations, building on the foundation of the Spark multimode reader platform. It combines sophisticated imaging with proven multimode reader capabilities, allowing you to define new approaches to your research and obtain robust orthogonal data faster than ever before.

Learn more about the detection features of the Spark Cyto

More throughput for your experiments

Spark Cyto can be extended with an automated multi-plate cell incubator or integrated in Tecan’s fully automated workflow solutions.

Learn more how to scale up your research

More experimental control

Designed to handle a broad range of common cytometry applications, Spark Cyto gives you a new level of experimental control without compromising on ease of use and convenience. Five predefined methods for common cytometry applications offer a straightforward approach to image acquisition and analysis, complemented by additional features such as ‘user-defined’ parameters and Real Time Experimental Control (REC™), making it possible for you to unlock new application possibilities

Learn more about SparkControl™ and Image Analyzer™

Multi-label live-cell cytometry characterizes your cells automatically

Spark Cyto capabilities*


  • 3Dai spheroid and organoid analysis
  • Label-free cell counting
  • Nuclei counting
  • Transfection efficiency
  • Cell viability
  • Apoptosis
  • Confluence assessment
  • Cell migration and wound healing
  • ELISAs
  • Low-volume DNA/RNA quantification
  • Nucleic acid labeling efficiency
  • Protein quantification
  • Reporter gene assays
  • HTRF®, DELFIA®, LanthaScreen®
  • Transcreener®
  • DLR®
  • BRET – including NanoBRET®
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Detection modes

  • Fluorescence imaging (blue, green, red, far red)
  • Bright field imaging
  • Digital phase contrast-imaging
  • Absorbance – incl. UV/VIS
  • Fluorescence top and bottom
  • Time resolved fluorescence (TRF)
  • Full spectral scanning capability for all measurement modes
  • FRET
  • Fluorescence polarization (FP)
  • Luminescence – glow, flash, multicolor, scanning
  • AlphaScreen®, AlphaLISA® and AlphaPlex®
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Additional features

  • Reagent dispensers with heater and stirrer
  • Humidity Cassette
  • NanoQuant Plate™
  • QC tools for IQ/OQ services
  • Spark-Stack™ microplate stacker
  • Multi-plate automated cell incubator extension

* Capabilities depend on the Spark Cyto configuration.

Tab 02 / Technology

Optics engineered for speed and clarity

A dedicated optical set-up for live-cell cytometry in microplates

The Spark Cyto fluorescence imaging module is designed to deliver crisp images with minimal user intervention. Using three different objectives, five LEDs (bright field and fluorescence excitation), a multiband filter set, and a 12-bit CMOS camera, Spark Cyto eliminates pixel shift and delivers high quality images in a flash.

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Schematic diagram of imager module. (1) LED for bright field; (2) microplate with sample; (3) objective; (4) multiband filter set; (5) LEDs and excitation filters for fluorescence; (6) Autofocus unit; (7) reflection mirror; (8) CMOS camera.

Built-in objective changer with 2x, 4x and 10x objectives.

Hassle-free selection of objectives and channels

Spark Cyto incorporates three objectives that can easily be selected on the fly in the SparkControl™ software:

  • 2x, numeric aperture =0.08
  • 4x, numeric aperture =0.13
  • 10x, numeric aperture =0.30

It also offers five LED channels to help meet the needs of your assay, without requiring manual user intervention.

  • Blue (381-450 nm)
  • Green (461-530 nm)
  • Red (543-611 nm)
  • Far red (626-800 nm)
  • Bright field and digital phase contrast
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Composite images without the pixel shift

The optical system requires no movement of the optics or plate transport to image a well with any combination of the five available channels. This eliminates pixel shifts and results in remarkable image quality and acquisition speed, especially when using two or more channels.

Autofocus enabled – stay focused on your research

Spark Cyto uses a patented LED-based autofocus system to deliver accurate images without compromising on scanning time. The autofocus system projects an extended grid pattern onto the sample surface, which minimizes the impact of potential distortions from isolated impurities, and then uses that as a guide to find the optimal focus for your samples. Simple, fast and effective autofocus that comes as standard on every instrument so that you never miss an image.

Live Viewer™

The Live Viewer software application enables the Spark Cyto to be used as an image cytometer, allowing cells to be monitored in real time. All optical channels and magnification levels are available in the Live Viewer, which can be operated from the software’s dashboard screen as a dedicated application, or from the Method Editor interface, to optimize parameters such as the focus offset or crosstalk between fluorescence channels before starting a measurement.

Analyze the whole picture

Whole well view of a 96-well plate recorded with the 2x objective; HeLa cells were stained with calcein AM and propidium iodide, and analyzed with the cell viability application. The whole well image reveals an area within the well where the local cytoxicity is significantly higher, which can point to effects related to compound dispensing.

Whole well imaging of 96- and 384-well plates

Spark Cyto allows you to image a whole-well in a 96- or 384-well plate with one single picture, giving you insight into every cell in every well. Using a proprietary patented wide field of view technology Spark Cyto obtains high quality whole-well images, without suffering from stitching artifacts or edge-to-edge image contrast. This gives you a complete picture of your cell population in less time, allowing you to drive your research in new directions.

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Single image of a whole well from a 96-well plate. No tiling or edge-to-edge optical distortion leads to superior results when analyzing cell populations.

One single image can tell the whole story

Many imaging systems advertise the capability to acquire high quality images of an entire well, but most lack the ability to do this effectively. These systems create composite images, using tiling or stitching, which misrepresent the cell population and suffer from distortion due to fluid meniscus shadows on the well borders.

Spark Cyto captures the whole well (96- and 384-well plates) with a single image giving you a real picture of your research.

Spark Cyto uses a patented approach combining image acquisition with the 2x (96-well plates) or 4x (384-well plates) objective with a large camera chip and advanced imaging algorithms to give you accurate results with one image.

  • 96-well plate – less than 12 minutes
  • 384-well plate – less than 45 minutes
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Imaging pattern for a whole well, 24-well recording using the 4x objective.

Whole-well imaging in other plate formats

Primary cell lines tend to show better growth dynamics in 6- to 48-well plates. Spark Cyto allows whole well imaging for these applications, based on the automatic generation of composite images of multiple single image tiles.

Spark Cyto is optimized for Tecan’s cell culture plates and this combination guarantees the best possible image quality for your cell-based assays.

Tab 03 / Applications

Five predefined applications, even more possibilities

Intuitive image acquisition and analysis methods

Spark Cyto takes a user-friendly approach to the most common cytometry applications:

  • 3Dai spheroid and organoid analysis
  • Label-free cell counting
  • Confluence
  • Nuclei counting
  • Transfection efficiency
  • Cell viability
  • Cell death

For all other applications simply select the ‘user-defined’ option to easily set up your own method in SparkControl™ or use the ‘image-only’ feature to export your files to a third-party image analysis software. This gives you complete flexibility in meeting your respective assay requirements.

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3Dai imaging features

Imaging of 3D cell culture objects is supported by AI-based segmentation of 3D structures, e.g., spheroids or organoids. Object-based data like area, eccentricity, diameter, and fluorescence intensity are derived from the segmentation mask and can be used to gate and analyze 3D objects. The Z- stacking function allows the acquisition of multiple layers in the Z-dimension and the automated creation of a projected image of these Z-layers, which serves as the basis for the AI-based segmentation.

Whole well image of a 96-well plate showing HeLa cells with bright field cell counting yellow overlay, acquired with the 4x objective.​

Label-free cell counting​

This feature offers a fast and reliable way to count cells in bright field images directly, without the need of cellular dyes.

Using a deep learning algorithm, a yellow cross is overlaid on identified cells for easy visualization, and the total cell count is displayed for each well.​

Whole well image from 96-well plate, acquired with the 2x objective; NNNHDF cells with confluence evaluation mask.


Use the bright field imaging channel to provide a quick overview of a well’s cell density. Cell confluence is calculated automatically by the software and displayed as a yellow overlay for easy visual confirmation. In addition, you can customize the roughness factor, a qualitative measurement, as a simple indicator of cell death.

Whole 384-well image acquired with the 4x objective; CHO cells with nuclei couting object mask.

Nuclei counting

Optimized for Hoechst 33342, this function provides an easy method for cell counting on the Spark Cyto, using any blue fluorescent dye with nuclear DNA binding capabilities.

Centered image of CHO cells cultured in a 96-well plate, acquired with the 4x objective, showing an overlay of the blue and green channels.

Transfection efficiency

This application is designed to automatically determine the transfection rate for cells containing green fluorescent protein (GFP), a widely used reporter for gene expression, and counter-stained with Hoechst 33342 blue nuclei dye. The green and blue images are overlaid and analyzed to determine the resulting transfection efficiency in the cell population.

Centered image of HeLa cells cultured in a 24-well plate, acquired with the 10x objective, showing an overlay of the bright field, green and red channels.

Cell viability

Spark Cyto’s preset cell viability application relies on a common double staining approach to discriminate between live (green) and dead (red) cells in a population. Using two fluorescent dyes, suach as calcein AM (live cells) and propidium iodide (dead cells), you can image and analyze your population in minutes.

Image of A431 cells cultured in a 96-well plate, acquired with the 10x objective, showing an overlay of the blue, green and red channels.

Cell death

The detection of, and discrimination between, apoptosis and necrosis can be accomplished by differential staining of markers characteristic of the relevant type of cell death, for example, with Hoechst 33342, propidium iodide and Annexin V-FITC.

Hoechst 33342 (blue) – nuclei stain

Propidium iodide (red) – necrotic cell stain

Annexin V-FITC / Alexa Fluor® 488 (green) – binds to the early apoptosis marker phosphatidylserine

Using a proprietary algorithm, the software can uniquely identify three object classes:

  • Blue objects – cell nuclei
  • Blue/red objects – necrotic cells (for live:dead cell ratio)
  • Blue/red/green objects – apoptotic cells (for apoptotic:necrotic cell ratio)
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Multi-Color analysis

Spark Cyto’s easy to use Multi-Color application is ideal for counting and analyzing cells with multiple labels, using a fluorescence marker for the nucleus and up to two additional labels to automatically characterize your cells.


Even more possibilities

Spark Cyto’s Method Editor offers two unique options for researchers looking to customize their assay:

  • User-defined protocols – set up specific imaging protocols for automated image acquisition and analysis
  • Imaging only – allowing acquisition and export of files to any third-party image analysis software, such as Image J or CellProfiler.

Real Time Experimental Control (REC™) – never miss a critical biological event!

REC grants you the ability to create new experimental workflows in your lab. Combining standard detection technologies, imaging capabilities and additional unique features, such as integrated humidity and environmental controls, REC unlocks new research possibilities.

REC uses all of these features to enable set-up of workflows that ensure you never miss a critical biological event – without chaining you to the bench.

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Complete environmental control comes as standard

Spark Cyto comes equipped with a unique set of environmental features for controlled measurement conditions in your microplate:

  • Uniform temperature control (up to 42 °C)
  • Dynamic gas control (CO2 and O2)
  • Humidity control via Lid Lifter™ and Humidity Cassette

In combination, these features allow you to maintain a stable environment for your assays, effectively eliminating the risk temperature fluctuations or evaporation could pose to your results. Spark Cyto is the only instrument to put these features right at your fingertips.

All of these functions are easily programmed and controlled with SparkControl software.

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Humidity control for optimal evaporation protection

Maintaining humidity levels of 95 % or higher is essential for unimpaired cell viability and growth, and miminizing evaporation is essential for maintaining consistent concentrations during long-term assays. Spark’s patented Humidity Cassette is a cost-effective solution to minimize evaporation.

Lid Lifter

Spark‘s integrated and patented lid lifting function creates new workflow possibilities, reducing the risk of sample contamination and helping to establish an ideal environment for long-term kinetic assays. Whether you want to dispense reagents without the need for manual intervention or maintain optimal environmental conditions without compromising evaporation protection, Spark Cyto is the only reader to offer this benefit.

Software designed for long-term studies

Spark Control combines automation of long-term kinetic assays with advanced features such as kinetic conditioning and remote monitoring, providing a hands-off solution for complex experimental set-ups.

Cell-based kinetic experiments require specific actions to be performed once a certain signal threshold is reached. Multimode readers without conditional kinetics make these studies cumbersome and potentially error-prone. SparkControl’s kinetic conditioning feature allows you to program the system to automatically perform any required action as soon as a predefined signal is reached, enabling all the assay workflows you can think of.

Tab 04 / Software

Spark Control™ – powerful control in an intuitive package

SparkControl, best-in-class user-interface software, now includes a dedicated imaging stripe specifically for Spark Cyto. The imaging stripe can be combined with any existing stripe in SparkControl, making the creation of multiplex assays straightforward and effortless. The new stripe is streamlined to give the operator complete control of all the parameters associated with their experiment, without burdening them with superfluous information and wasting valuable time.

The powerful reader control software of the Spark Cyto, SparkControl besides the 2D imaging stripe, includes a new stripe to set up and optimize automated 3D cell culture imaging protocols. The stripes can be combined with any other programming stripe of the conventional detection methods to allow easy multiplexing. The operator can select the objective and imaging channels of choice, define the field of view and control all parameters associated with the measurement. A quick link to the microscope-like Live Viewer mode gives immediate access to the image acquisition settings.

SparkControl software offers a live viewer mode that turns the reader into a digital microscope, allowing manual inspection of cells, as well as optimization of focus levels, exposure times and LED intensities.

Image Analyzer™ – robust and accessible analysis tool

Images acquired with the Spark Cyto can be automatically processed with Image Analyzer, Tecan’s proprietary imaging software package, offering easy data analysis for object segmentation, counting and gating.

Spark Cyto's Image Analyzer allows easy data analysis for object segmentation, gating, object counting in brightfield and fluorescence images. For 3D object functions it allows the re- analysis of Z-projection images from existing Z-stacks.

Using the Image Analyzer, parameters such as brightness and contrast can be individually adjusted for each channel. Furthermore, overlay images can be generated by integrating channels and object masks from the analyzed parameters.

Tab 05 / Configurations

Configurations to meet your applications

Spark Cyto is available in five specialized configurations, which differ only in their detection modalities. This means that no matter the configuration of your Spark Cyto, you have a fully equipped system ready for real-time live cell imaging cytometry.

Every configuration of the Spark Cyto includes the following features for real-time cytometry, truly redefining what is standard for an imaging microplate reader:

  • Lid Lifter™
  • Integrated gas control (CO2/O2)
  • Heating
  • LED-based autofocus
  • Objectives (2x, 4x and 10x)
  • 5-LED excitation, 4 color channels
  • Digital phase contrast
  • SparkControl™ software
  • Image Analyzer™ software
  • Instrument control unit

All five configurations can be optionally combined with additional features - see overview.

Tab 06 / Accessories


Automation for higher throughput

Tecan’s Spark Motion concept enables walkaway automation for live cell experiments on up to 40 plates.

  • Automated cell incubation and analysis of up to 40 plates
  • Multiplexed kinetic growth analysis (eg. luminescence and imaging) increases reproducibility
  • Patented lid-lifting technology in Spark Cyto protects plates outside the incubator, saving costs for expensive safety cabinets
  • Expandable with additional instruments
Interested in complete workflow automation?

Learn more about Tecan’s fully automated solutions

Read more

Spark-Stack™ microplate stacker*

Spark-Stack is a compact, automated plate stacker for integration with Spark readers. It provides reliable processing of up to 50 plates per run.

Injectors with heating and stirring

Integrated injectors permit automated reagent dispensing, ideal for high sensitivity applications such as flash luminescence. An attachable heater/stirrer unit reduces the risk of precipitation.

Humidity Cassette

Built-in evaporation protection enhances live cell kinetic assays, minimizing edge effects for better uniformity and more reliable data.

A patented Humidity Cassette reduces evaporation in standard microplates, enabling long-term live kinetic studies without the need to switch to dedicated and costly microplate types – simply use your current, validated plates. Benefits include:

  • Reduction in evaporation during your assay
  • Use your existing plates – no need for expensive special plates
  • Better data quality – consistant results
  • Reduced edge effects – ideal for long-term kinetics
  • Stable conditions – better cell viability
Read more

QC Tools and IQ/OQ services

Tecan’s quality control packages help you meet all regulatory requirements in an efficient, cost-effective way.

Read more

* Spark-Stack microplate stacker supports all read modes without imaging.

Tab 07 / Webinars

Spark Cyto webinars

Combining live cell imaging with industry-leading detection technologies, you now have the ability to unite qualitative and quantitative information into unique multiparameter data sets.

These educational webinars explain how scientists advance their research with enabling cell analysis techniques.

Never miss a critical biological event with improved whole-well imaging

Dr. Stefan Hasenöder, Sirion Biotek

This webinar offers you insights into how to improve the robustness of your live cell assays, addressing and overcoming some specific causes of reproducibility problems.

New methods to better predict patient drug responses in cancer organoids using a multimode imaging plate reader

Asst. Prof. Dannielle Engle, SALK institute

Pancreatic cancer is a deadly malignancy with few treatment options. This webinar highlights how dynamic measurements of confluence in patient-derived organoid models of pancreatic disease can predict treatment responses.

Semi-automated multiplex assays for monitoring therapy response in patient-derived 3D organoids

Dr Christoph Deben, University of Antwerp

This webinar explains how you can set up experimental workflows and multiplex analysis techniques to monitor drug responses in real time.

Streamlining high-content cytotoxicity assay development

Dr. Christopher Wolff, FMP

If you are looking for a fast, uncomplicated and effective way of developing cytotoxicity cell painting assays, you won’t want to miss this webinar. Join Christopher Wolff from FMP Berlin and Christian Oberdanner from Tecan in an informative illustration of the precise and sensitive characterization of cytotoxicity based on multicolored imaging with Spark Cyto.

Advantages of tumor-specific 3D models

Dr. Julia Kirshner, zPREDICTA

Find out about the benefits of using 3D culture models in oncology drug discovery. This webinar will focus on the importance of the tumor micro-environment and tissue-associated extracellular matrix (ECM) for obtaining accurate drug response data, using the novel analytical capabilities of the Spark® Cyto multimode imaging plate reader.

Harnessing HT metabolomics and phenotypic profiling for cancer research

Dr. Mattia Zampieri, ETH Zürich

Metabolic profiling of cancer cell line collections has become an invaluable tool in the study of disease etiology and drug modes of action, as well as for selecting personalized treatments. However, its scale is limited by time-consuming sampling and complex measurement procedures.

Novel cell models for rapid screening of SARS-CoV-2 infection studies

Dr. Marek Widera, Alexander Wilhelm

To decipher the pathology of COVID- 19, in vitro cell culture models that can physiologically mimic the viral replication cycle are required. This webinar introduces the A549-AT cell line, generated to meet this need and enable rapid and sensitive monitoring of SARS-CoV-2 replication and facilitating the characterization of viral variants.

Live-Cell, Real-Time Cell Imaging and Analysis for Drug Discovery

Understanding when and how cells die following drug treatment is critical in the drug discovery and characterization process. However, traditional cell-based experiments that rely on endpoint data collection lack the details to answer these questions. Learn how to make more informed decisions on drug candidates with live-cell imaging and kinetic data using high-throughput amenable assays and instruments.

Gaining insights into DNA damage response (DDR) in cancer cells with image-based quantification

Maintenance of genome integrity is essential for the prevention of mutations and cellular transformation, which can give rise to cancer. Find out how the Spark® Cyto was used to gain insights into DNA repair dynamics in living cells and study the DDR upon treatment with chemotherapeutic agents in fixed cells.

Advantages of new AI-based stain-free cell analysis for cancer research

Dr. Roland Zauner, EB House Austria

In various cell-based assays, the number of cells is either determined as the primary measurement, as in proliferation assays, or used as a reference to normalize readouts. Find out how the new label-free cell segmentation tool can be applied in anti-cancer drug screenings and how research into the genetic skin disease epidermolysis bullosa will benefit from this development.

Whole-Well Brightfield Cell Counting with the Latest in Microplate Imaging Technology

Dr. Christopher Wolff, FMP

Label-free imaging and analysis of cells using the brightfield imaging channel is a non-invasive, non-toxic alternative to fluorescent microscopy, but is challenging using conventional microscopes and automated imaging systems. Find out how cutting-edge neural network-based algorithm technology enables rapid, label-free cell counting, creating new opportunities for researchers to conduct live cell experiments and cell analysis.

Automated, multiplexed, and reproducible analysis of 3D cell-based research using Spark Cyto.

The evolution of drug discovery requires physiologically relevant models. Rethink the 2D cell models of the last generation and attain reproducibility at scale with 3D organoids and spheroids from 6- to 384- well plates.

Tab 08 / Literature


The Spark Cyto is a multimode reader platform equipped with a highly sophisticated fluorescence imaging module for real-time cytometry.

Application Guide


Applications Notes

Predictive Oncology is a science-driven company offering solutions that introduce human diversity at an earlier stage of the drug discovery process. The company has developed a 3D culture model, r-Breast, to replicate the conditions in the mammary tissue and recreate the tumor microenvironment, including the epithelial and stromal niches. This provides a physiologically-relevant approach to evaluating new drugs or drug combinations, while also accounting for species-specific differences in the extracellular matrix composition. This application note describes a study of two different co-culture systems using breast fibroblasts and T-cells. First, fibroblasts were cocultured with breast cancer cells to assess the impact on drug sensitivity. Second, the effect of programmed cell death protein 1 (PD-1) drug treatment on the viability of breast cancer cells in the presence of T-cells was investigated. Cell viability was assessed using the r-Breast model with propidium iodide-based assays and CellTiter-Glo® luminescence analysis (Promega), efficiently evaluating the results using the Spark Cyto multimode reader.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has become a global concern, due to its rapid spread. The study, published by the University of Frankfurt, describes the use of the Spark Cyto for the development of a cellular infection model that enables high throughput SARS-CoV-2 experiments and live cell imaging. The automated, non-invasive optical readout included assessment of confluency, roughness factor and fluorescence measurement. The data further highlights the use of the cell line for screening for antiviral compounds as well as for investigating the efficacy of neutralizing antibodies against different SARS-CoV-2 variants.

This application note systematically defines experimental parameters to enable live cell nuclear staining with minimal cytotoxicity during repeated exposure and continuous fluorescence imaging in the Spark Cyto, as well as robust automated cell segmentation with the SparkControl™ and Image Analyzer™ software.

The optimized procedure was used in a multiplexed, three-color assay to detect and distinguish early and late stages of apoptotic cell death, opening the door for dynamic long-term phenotype tracking.

Alzheimer’s disease (AD) and vascular dementia (VaD) are the two most common types of dementia, and their incidence is increasing year by year. They affect the health of the elderly, and cause a huge burden on society, with no effective treatments currently available.

In vitro research is the first step in drug evaluation, and cellular immunofluorescence can be used to more clearly define the role of drugs, provide more comprehensive and accurate information for downstream drug development.

This study uses the Spark Cyto – an innovative multimode reader with cell imaging capabilities – to evaluate the neuroprotective effects of miR-23b-3p (miRNA) and tilianin (compound) on stable transfected cell lines and human primary neurons. The results in this application note are based on a peer-reviewed article, published in the journal Oxidative Medicine and Cellular Longevity in August,2021, with the title: Tilianin Ameliorates Cognitive Dysfunction and Neuronal Damage in Rats with Vascular Dementia via p-CaMKII/ERK/CREB and ox-CaMKII-Dependent MAPK/NF-κB Pathways.

Scientific innovation is dependent on the power of observations, and the strength of the conclusions drawn from those observations. In in vitro biology, a majority of physiologically relevant outcomes adhere to dose- and exposure-dependent factors. Unfortunately, data collection for traditional cell-based experiments often occurs at arbitrary but convenient endpoints, and using inadequate or poorly informative tools. The resulting data typically lack the appropriate kinetic resolution to fully answer details of the cause-and-effect relationship. Because biological pathways and processes are already inherently complex, a more efficient screening paradigm is required, allowing scientists to examine the important parameters of ‘when’ and ‘how’ to better characterize a given response. This application note seeks to provide a practical demonstration of how real-time assays and a plate reader with bright field and fluorescence imaging functionality can work in unison to reveal cell- and compound-specific features of an apoptotic response.

This application note describes the outcome of an experiment comparing the Opera Phenix with the Spark Cyto for the detection of apoptotic cells. For this study, Hoechst 33342 was used as a ‘blue’ nucleic counter stain, with TMRM and YO-PRO-1 as the two secondary mask signals. Please note that findings presented here do not necessarily reflect the general performance difference between the two systems.

Workflow automation with the Spark Cyto multimode reader.

Automated monitoring of the reporter activity of hfob.1.19 cells following induction of osteogenic differentiation by temperature shift

While 3D cultures are superior to the 2D culture approaches in terms of physiological tissue organization and providing robust prediction of clinical outcomes, these techniques are often more expensive and time consuming to perform compared to the standard 2D protocols. Performing multiplexed assays with various readouts is therefore beneficial in order to gain the most information from each experiment. Here, we describe a multiplexed approach combining sequential imaging based LIVE/DEAD cell viability assays (Thermo Fisher Scientific) and CellTiter Glo luminescence analysis (Promega) using the Tecan Spark Cyto with r-Breast and r-mBreast models.

The genomic integrity of mammalian cells is constantly challenged by DNA damage arising from both endogenous and exogenous sources. Complex DNA repair pathways have evolved to deal with specific types of DNA lesions. An efficient DNA damage response (DDR) requires immediate detection and repair of the damage.

In addition, cell cycle checkpoints need to be activated to allow enough time for repair, prevent further damage through collision of the replication and transcription machinery with unrepaired lesions, and ensure that lesions are not passed on to daughter cells.

While impaired DDR can lead to serious diseases like cancer, it also presents an opportunity for therapeutic interventions exploiting these repair defects. 1 DNA repair is a highly dynamic process involving distinct and well-coordinated steps, and should therefore ideally be studied in living cells.2 However, a lot of post-translational modifications essential for the DDR can only be analyzed in fixed cells.

Highly aggressive cutaneous squamous cell carcinomas (SCCs) occur particularly frequently and early in patients suffering from the rare genetic skin disease recessive dystrophic epidermolysis bullosa (RDEB), causing a high mortality rate...

Predictive Oncology’s r-Tube 3D model mimics the niches in which endothelial cells reside within tissues, providing a more physiologically accurate platform. This allows the effectiveness of novel drugs or drug combinations to be evaluated in a setting that captures the species-specific differences in ECM composition. In this manner, the r-Tube 3D system offers a valuable tool for studying cancer and contributing to the development of improved therapeutic strategies, by incorporating the microenvironment into the culture model. The Spark Cyto has been widely used for bright field and fluorescence imaging in 3D models and has proven suitable for imaging thick samples with cells embedded in a 3D matrix. A multiplex approach using the r-Tube 3D model to investigate eribulin and ramucirumab treatment was therefore performed using the Spark® Cyto with sequential bright field imaging and CellTiter- Glo® luminescence analysis (Promega).

The essence of 3D liver tissue reconstruction lies in creating functional liver tissue outside of the body, mimicking the native ECM and microenvironment. This approach offers several advantages. First, it enables the study of liver pathophysiology and disease mechanisms, leading to better understanding of novel therapeutic targets. From a regenerative medicine standpoint, 3D liver tissue reconstruction offers patient-specific solutions; patient-derived tissues can be created using primary liver cells, minimizing the risk of immune rejection and eliminating the reliance on donor organs. These engineered liver tissues hold great promise for transplantation and tailored treatment plans. Furthermore, 3D liver tissue models provide an ethical and clinically relevant platform for drug testing, reducing the need for animal testing. This application note describes the combination of Tecan’s Spark Cyto reader with Predictive Oncology’s r-liver-tox set-up to perform multiplexing investigations, including live/dead cell analysis, detection of intracellular production of reactive oxygen species (ROS) and the immunostaining of primary human hepatocytes.

Predictive Oncology has developed a three-dimensional (3D) model called the Reconstructed Pancreas (r-Pancreas), which attempts to replicate the pancreatic tumor microenvironment. It successfully recreates the conditions found in pancreatic tissue – including both the epithelial and stromal niches – offering a more physiologically relevant approach to assess new drugs, or combinations of drugs. As described in this Application note, this research model was used in combination with the Spark Cyto multimode reader to investigate the efficacy of gemcitabine, a therapeutic agent against pancreatic tumor cells. To enhance the physiological relevance of the r-Pancreas model, a collagen-rich capsule was introduced, as human pancreatic tumors are known to possess an outer layer of stiff ECM, acting as a physical barrier that hinders drug penetration. Moreover, by incorporating a co-culture of primary activated pancreatic stromal cells and pancreatic tumor cell lines, it was possible to create a more comprehensive 3D model that allows testing of potential therapeutic agents within the pancreatic tumor microenvironment. In this model, pancreatic tumor cells were co-cultured with human pancreatic fibroblasts.

Streamlining 3D cell culture-based drug response profiling with the Fluent® Automation Workstation, D300 Digital Dispenser, and Spark® Cyto multimode reader.

The development of treatments for snake bite induced vascular toxicity relies on robust, quantitative, and relevant in vitro models of blood vessels. Organ-on-a-chip is an emerging technology that uses miniaturized components to create a three-dimensional culture of human cells grown in a microfluidic system. In contrast to cells grown in 2D, blood vessels grown in microfluidic channels enable the inclusion of several important physiological parameters during cell culture, such as 3D tubular morphology, fluid perfusion, inclusion of extracellular matrix (ECM) and exposure to biochemical gradients. Real-time assays combining organ-on-a-chip platforms with multimode readers would enable direct monitoring of vascular barriers, viability and morphology with a sufficient temporal resolution to capture the fast effects of potent snake venom toxins. This application note describes the use of the Spark Cyto to multiplex readouts and comprehensively assess the potential of different snake venoms to cause vascular toxicity of a human blood vessel model grown in an organ-on-a-chip platform.

Technical Notes

Analyzing the dose-response relationship using cell-based assays is essential to understanding a drug’s efficacy. However, fixing the appropriate dose range and time point is often challenging. The D300e Digital Dispenser enables accurate delivery of compounds at picoliter levels, spanning a wide dose range – for example, from 0.2 nM to 1 μM – in just a few minutes. This can be combined with the live-cell imaging and full cell incubation capabilities of the Spark Cyto multimode reader to monitor cell proliferation under standard culture conditions (37 ºC, 5 % CO2). This technical note uses the D300e and Spark Cyto to comprehensively capture an anti-cancer drug’s efficacy profile, tracking cell viability with nine doses at 25 time points using luminescence-based and bright field imaging technologies simultaneously.

This Technical Note briefly describes how to use Image Analyzer for the optimization of confluence assessments in bright field images, as well as for object segmentation and counting in fluorescence images.

This application note describes a semi-high throughput approach for the analysis of intracellular Ca2+ and cellular Ca2+ uptake using the Spark Cyto’s multicolor fluorescence imaging and the Image Analyzer™ software’s integrated Voronoi analysis function. Treatment with the calcium ionophore ionomycin was used to increase intracellular Ca2+ concentrations, and Fluo-4 was used in combination with the nuclear counterstain Hoechst 33342 for measurement quantification.

This technical note describes the capabilities of the Spark Cyto to normalize a cellular signal to the cell number, or cell mass, per well. This dramatically improves the overall data quality of the respective cell-based assay and, finally, leads to an improved reproducibility of cell-based experiments and more efficiency in the lab.

Fluorescence imaging using the spark cyto – acquisition in a snap using sparkcontrol™

Accurate and automated assessment of cell density using bright field and fluorescence imaging…

Automating visualization and quantification of protein expression in microplates…

Discriminating viable, apoptotic and necrotic cells.

Quantification of life: Dead less ratio with the Spark Cyto imaging cytometer.

SparkControl™ offers a wide selection of predefined plate definition files (.pdfx), and it is crucial to use the correct plate definition for imaging applications to avoid autofocus errors. Alternatively, the Plate Geometry Editor can be used to modify existing files to suit unlisted plate formats.

This Technical Note describes the use of the Plate Geometry Editor in SparkControl™ and points out the most important aspects for the successful creation of pdfx files.

A non-invasive way to accurately count cells in brightfield using a deep learning algorithm.

Cell line development plays a crucial role in modern biological research and drug development. Clonal lines derived from a single cell can provide a reproducible and stable platform for drug discovery, studying biological processes and producing recombinant proteins. Genetic engineering and genome editing techniques have enabled the creation of cell lines with specific mutations or gene knockouts, allowing researchers to investigate the function of individual genes and pathways. Additionally, cell lines are used in the production of biologics, such as monoclonal antibodies and recombinant proteins, making the development of new and improved cell lines a critical area of research in both academia and industry. This technical note describes a method for using the Spark Cyto multimode reader to perform whole-well imaging of single mammalian cell clones in 96-well plates, in order to identify optimal clones. Both the Spark and Spark Cyto are capable of verifying monoclonality, however the Spark Cyto offers a faster readout and improved image quality compared to the Spark’s basic imaging functionality, providing higher throughput clone identification.

This technical note describes the use of the Uno Single Cell Dispenser to isolate single cells. This instrument harnesses microfluidic digital dispensing technology to gently isolate viable cells for various downstream applications, including cell line development.

The viability of the single cell post dispensing was assessed by addition of a fluorescent dye, that bound to the DNA of cells with impaired membrane integrity, using whole-well fluorescence imaging functions of the Spark Cyto.

Cell outgrowth rate was determined 10 days after single cell isolation with the Uno, using whole-well brightfield imaging. Using Uno and Spark Cyto together, enables simple single cell isolation and determination of monoclonal cell outgrowth for variety of downstream application.

Recent advancements in cell-based research strategies have revolutionized drug discovery, disease modeling, tissue engineering, and personalized medicine. The shift from traditional 2D monolayer culture to three-dimensional (3D) cell culture systems has opened new avenues for more realistic representation of complex tissues within the human body. Particularly the use of multicellular 3D spheroids is showcased here. These spheroids represent a significant advancement in cell-based assays. In this note, the generation of spheroids in Corning 96-well, round bottom ULA microplates and subsequent monitoring and analysis of spheroid growth using the Spark Cyto multimode reader is presented. The integration of a plate washer (HydroSpeed™) further optimized the workflow, enhancing automation, standardization, and reproducibility, while reducing human errors. These advancements represent a substantial step forward in enhancing our understanding of cellular behaviors and developing more realistic in vitro models for various applications.


Bringing time and cost advantages to labs…

How to guide

Cell-based assays have a variety of practical uses within scientific research and are an integral component of many scientists’ workflows from testing tumor resistance for cancer research to helping identify novel therapeutic compounds in drug discovery.

LSIA2019 logo Silver

Silver winner of the Bioinformatics Life Science Industry Award 2019 in the category:
Innovation – Best New Product Cell Biology.


The Spark Cyto can be an excellent support in any cell biology laboratory.

Fabio Gasparri, Nerviano Medical Sciences on SelectScience

Brochure and Typical perfomance values

The Spark Cyto is a multimode reader platform equipped with a highly sophisticated fluorescence imaging module for real-time cytometry.

Read the Spark Cyto brochure
See the typical performance values for Spark Cyto

Automation for higher throughput

Tecan’s Spark Motion concept enables walkaway automation for live cell experiments on up to 40 plates.

  • Automated cell incubation and analysis of up to 40 plates
  • Multiplexed kinetic growth analysis (eg. luminescence and imaging) increases reproducibility
  • Patented lid-lifting technology in Spark Cyto protects plates outside the incubator, saving costs for expensive safety cabinets
  • Expandable with additional instruments
Interested in complete workflow automation?

Learn more about Tecan’s fully automated solutions

Read more

Automated, multiplexed, and reproducible analysis of 3D cell-based research using Spark Cyto.

The evolution of drug discovery requires physiologically relevant models. Rethink the 2D cell models of the last generation and attain reproducibility at scale with 3D organoids and spheroids from 6- to 384- well plates.

Spark Cyto is for research use only.