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The Beginning of a New Era: New Microfluidics Technology Advances Cytometry

The innovative use of fiber optics in flow cytometry ushers in a new era via a rugged, ultra-compact, lightweight platform for daily, routine flow cytometry applications. This simplified and cost-effective personal cytometry platform developed by handyem is vibration-proof and can be easily transported from laboratory to laboratory. The personal cytometry (HPC) platform is suitable for use under a biological hood, in a glove box, or for fieldwork.

This paper describes how the use of fiber optics facilitated the vision to simplify and generalize the use of flow cytometry.

Introduction: The Advent of Personal Cytometers


Flow cytometry is recognized as an especially useful technology for analyzing cells and particles. In practice, most researchers rely on core laboratories which possess state-of-the-art instruments and highly-trained staff to help plan and carry out complex experiments.

These facilities use high throughput instruments to service routine applications and normally charge a fee based on instrument cost. As a matter of fact, high-end instruments are unnecessary for applications requiring the use of one to four colors. They are also too complex for non-expert cytometrists.

In recent years, technology evolutions have enabled the miniaturization of flow cytometers. Flow cytometry specialists observed that “technological milestones have already brought flow cytometry out of centralized core facilities while making it much more affordable and user friendly[1] . A new breed of smaller flow cytometers made “personal cytometers” possible. These personal cytometers are commonly available nowadays and offer many benefits to traditional and less traditional flow cytometry users.

However, these new cytometers are built with the laser and free-space optical architecture from the early instruments, making them hardly portable and equally sensitive to vibrations. Moreover, such cytometers require a similar level of care and maintenance as their legacy ancestors. This is where handyem made a step forward with the introduction of an innovative technology platform that integrates fiber optics, providing the ruggedness and portability that was missing in flow cytometry instrumentation.

We will now describe in general terms how traditional cytometry works and explain the novel technology, pointing out the differences between both.

Traditional Flow Cytometers: Hydrodynamic Focusing and Free-space Optics

Conventional flow cytometers use sheath fluid to suspend and hydrodynamically focus cells or particles in a liquid stream as they pass in front of a laser light source. Traditional hydrodynamic focusing requires precise laser beam alignment, adds bulk and complexity while commanding higher running costs. In addition, the interrogation area where the excitation laser hits the cells or particles can vary with the flow rate.

Figure 1: Hydrodynamic focusing

The critical component of a flow cytometer is the excitation laser. Its emission beam must hit a very small target (less than 20 µm) precisely and consistently to ensure that the cells or particles passing in front of it receive the same level of excitation, i.e. the same quantity of photons. Traditional flow cytometers use a combination of free space and bulk optical components such as lenses and mirrors held and aligned with precision using mechanical fixtures (Figure 2). The laser’s beam alignment must be assured which increases operation complexity and prevents potential portability of the instruments.

Figure 2: Optical architecture of traditional flow cytometers/free-space optics

Development of the HPC Platform

Founded in 2011 with the mission to simplify and generalize the use of cytometry, handyem’s goal is to provide non-flow cytometry experts with cell- and bead-based assay capabilities.

The idea of a cytometer based on fiber optics began in 2006 at the National Optics Institute (INO), a world-class Canadian center of expertise in optics and photonics. During the development process, this research raised interest from the Canadian Space Agency (CSA). The agency was looking for an instrument to monitor the health of astronauts on the International Space Station (ISS). The outcome was INO’s Microflow, the first flow cytometer to function in outer space (see sidebar).

Several patents were issued for a principle of measurement incorporating new micro-laser machining technology. INO’s flow cytometry patents were acquired by handyem to develop the HPC platform; the first commercial breed of fiber-optics-based flow cytometers.


Canadian astronaut Chris Hadfield demonstrated the Microflow, a miniaturized flow cytometer, in the microgravity environment of the International Space Station. The instrument supported immunopheno-typing as well as microbead-based multiplexed cytokine assays in the space environment and independently of gravity levels.[2]

Fiber Optics Architecture: Excitation and Collection Fibers

FiberFlowFluidicsTM, the innovative technology at the core of the HPC platform, uses fiber optics to guide the light beam from the laser to the small interrogation area to ensure consistent excitation of the cells or particles. The beam-delivery fiber, the forward scatter (FSC) and side scatter (SSC) collection fibers are bound in a monolithic assembly (Figure 3). The resulting flow cell is unsusceptible to vibrations and cannot be misaligned which allows the HPC instruments to be moved or carried without laser beam alignment concerns or adjustments by a specialized technician – a costly and time consuming operation.

Figure 3: FiberFlowFluidicsTM flow cell – monolithic fiber optics architecture

The sample crosses the micro-cavity in the rectangular collection fiber. The excitation fiber delivers the laser light to the flow cell and acts as a spatial filter, creating a very uniform beam that strikes the micro-cavity fluid channel (Figure 4). A fiber located on the opposite side of the flow cell collects the forward scatter light (FSC), while the collection fiber transports side scatter (SSC) and fluorescent light to the detection module – an arrangement of optical filters and detectors. A broadband mirror located at the end of the collection fiber reflects the side scattered and fluorescent light to increase the detection efficiency.

In the early 1840s, Daniel Colladon and Jacques Babinet first demonstrated guiding of light by refraction, the principle that makes fiber optics possible. An optical fiber is a flexible, transparent fiber made of glass or plastic that functions as a light-transmitting conduit.[3]

Figure 4: Light propagation in FiberFlowFluidicsTM flow cell

The thickness of the collection fiber allows for a longer optical path, much larger than the cell or particle, and the generation of a larger pulse width. Unlike traditional flow cytometers, the area measurement in the HPC platform is directly proportional to the height parameter. Furthermore, the use of unique signal-processing algorithms improves the signal-to-noise ratio and sensitivity.
FiberFlowFluidicsTM Allows Compact, Rugged, Micro-sheath and Sheathless instruments

Additionally, fiber optics enable ground-breaking microfluidic flow cell design that requires minimal - or no - volume of sheath fluid. Some advantages of the micro-sheath or sheathless fluidic systems include decreased instrument complexity and overall usage costs through the reduction of footprint and consumable expense (Figure 5).

Figure 5: Micro-sheath and sheathless flow cytometry

Sample Reuse with the HPC-Platform

The HPC-100 is the sheathless version of HPC platform. It uses a syringe pump to draw from the sample from a 1.5 mL microtube and direct it through the flow cell for cytometry measurement. The sample is temporarily stored in a small tubing loop (the buffer). The user can decide whether to recover the sample in the microtube or to send it to the waste bottle for disposal.

Furthermore, the flow cell and the detection module do not need to be mechanically aligned because the light signals are routed through fiber optics; hence the complex assembly of “mirrors and lens” in typical flow cytometers is not necessary. The laser and detection modules can be positioned to optimize the instrument design and reduce overall system dimensions (Figure 6).

Figure 6: HPC block diagram – contribution of fiber optics to compactness

In contrast with bulk free-space optical components used by conventional cytometers, the FiberFlowFluidicsTM technology integrates compact and flexible fiber optics which enables small packaging (Figure 7).
Figure 7: Ultra-compact and portable HPC instruments
Broad Dynamic Range and PMT Voltage Adjustment

The detection module includes an arrangement of filters and dichroic mirrors and photomultiplier tubes. The signals from the photomultiplier tubes (PMT) of the fluorescence channels are amplified and filtered before digitization. The highly sensitive electronic circuitry provides a dynamic range with more than six orders of magnitude. This allows to see the big picture in a flash without the hassle of gain or threshold adjustment. Optimization of the PMT voltages provides additional flexibility to avoid saturation or to increase the signal-to-noise ratio (SNR). The benefit: faster time to visualize measured data.

FCS file format

The Flow Cytometry Standard (FCS) is a file format supported by all flow cytometry instrument and analysis software vendors. Over the years, the standard has evolved through many revisions. FCS 3.1 is the current and most up-to-date version (introduced in 2010).[4]

Acquisition Software Stores Data in FCS Files for Easy Export

The HPC instrument operation is controlled by CytodyemTM Light data acquisition software. Its easy-to-use graphical user interface allows the user to scan samples, display up to 6 scatter plots and histograms, set gates, zoom and store the data in standard FCS file format (see sidebar). It also provides routine and microfluidics maintenance functions such as cleaning, rinsing, decontamination and many others.

Figure 8: Screenshot from CytodyemTM Light data acquisition software

The HPC Platform in a nutshell

The HPC platform provides a performance comparable to other commercially-available personal flow cytometers. Rugged and compact, the HPC instruments can be used for a large variety of biological applications including cell counting, apoptosis, cell cycle and DNA analysis, GFP transfection along with human and animal reproduction studies. The HPC platform is vibration-proof hence absolutely portable.


The HPC instruments can be configured with one or two excitation lasers (Blue/488 nm, Green/532 nm or Red/638 nm) and up to 4 fluorescence channels for a maximum of six measurement parameters. The FiberFlowFluidicsTM patented technology delivers simplicity at an affordable price, lowering the cost of ownership without any trade-off in performance.

The HPC personal cytometers are:

  • Compact and lightweight
  • Vibration proof and portable
  • Easy-to-use
  • Cost effective
  • Low-maintenance
  • High-performance

Recent technology evolutions have enabled the miniaturization of flow cytometers. The new breed of smaller flow cytometers made “personal cytometry” possible. However, these new cytometers are built with the same laser and free-space optical architecture, making them hardly portable and equally sensitive to vibrations. This paper showed how the introduction of an innovative technology platform that integrates fiber optics, provides the ruggedness and portability that was missing in this practice, and opens an array of new possibilities in flow cytometry.

The novel technology, named FiberFlowFluidicsTM, resulted in micro- sheath, ultra-compact, lightweight instruments, designed for daily use for the majority of common flow cytometry applications. The simplified and cost-effective HPC instruments are rugged and vibration-proof, and can be easily transported for use under a hood, in a glove box, in another laboratory, or for fieldwork. This new platform is a step forward in simplifying and generalizing the use of cytometry.


  1. Shapiro, H.P., The Evolution of Cytometers, Cytometry Part A, Wiley-Liss, Inc., 2004, 58:13-20.
  2. Dubeau-Laramée G, Rivière C, Jean I, Mermut O, Cohen L.  Microflow1, a sheathless fiber-optic flow cytometry biomedical platform: Demonstration onboard the International Space Station. Cytometry Part A. 2013 November; epub. DOI:10.1002/cyto.a.22427. PMID: 24339248.
  3. David R. Goff. Fiber Optic Video Transmission, 1st ed. Focal Press: Woburn, Massachusetts, 2003 and other private writings.
  4. International Society for Advancement of Cytometry (ISAC)

For Research Use Only. Not for use in diagnostic or therapeutic procedures


  1. (Shapiro, 2004)
  2. (Dubeau-Laramée, 2013)
  3. (Goff, 2003)
  4. International Society for Advancement of Cytometry (ISAC)