Microfluidics for Cell Biology
The Advantages of Flow Control for Cell biology and Microscopy
- Shear stress control
- Automation of multiple fluid delivery
- Long term perfusion
What are the advantages of microfluidics in biology?
The development of microfluidics has brought many advantages for biological research. Indeed, due to the miniaturization and the automation of the setups, the consumption of sample and reagent is largely decreased, the experiment times are considerably reduced and the global costs of the applications are minimized. Moreover, working with small quantities allows better carry out separations and detection with high resolution and sensitivity at reduced costs.
Finally, thanks to this new generation of techniques and equipments, new capabilities are offered for biology research since microfluidics allows to obtain more accurate results, to reduce detection limits and even to perform multiple analyses simultaneously.
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Main applications of microfluidics in cell biology
RNA and DNA Hybridization
For decades, in situ hybridization has been used with greater frequency to capture the localization, structure and expression of specific DNA and RNA sequences within tissues. The high sensitivity and specificity of this technique relies on the hybridization of labelled oligonucleotide probes to targeted DNA or RNA sequences within tissues. Chromosomal microdeletion, amplification, structure, translocation and expression can be easily detected.
Compared to PCR analysis, Northern Blotting or a DNA microarray performed on lysed cells, hybridization provides spatial information as specific RNA and DNA are detected within tissues. It has significantly improved gene mapping, cytogenetics and various diagnostic techniques (oncogenic, prenatal, viral infection, etc.).
Recently, with the use of microfluidic for cell biology, a temporal dimension was added to the technology, as it further evolved to be performed on living cells. The spatio-temporal expression, degradation and storage of RNA molecules can now be thoroughly investigated by real time imaging of the hybridization of labelled oligonucleotides in living cells.
Drug Combination and Long-Term Perfusion
In various therapeutic areas (glaucoma, vascular, HIV, oncology), a combination of drugs is typically found to be more effective for disease treatment. Oncology, in particular, has paved the way for this approach. The emergence of next-generation sequencing technology profiling has revealed the heterogeneity in many cancers at the origin of the differential response to treatment. As a result, therapeutic strategies are evolving towards multi-targeted drug combinations that effectively inhibit the cancer cells and block the emergence of drug resistance while selectively incurring minimal side effects on healthy cells.
Drug combination therapy is not straight forward as in many cases the results do not equal the sum of the parts. Cross-reactions are observed and fall into 5 categories: low risk & synergy, low risk & no synergy, caution, unsafe, and dangerous. The contribution and dosage of each active molecule should be closely investigated in terms of dose, frequency and duration to evaluate the efficacy of the drug combination.
Therefore, microfluidics for cell biology holds great promise in cancer diagnosis and also serves as an emerging tool for understanding cancer biology. Microfluidics can be a valuable tool in cancer investigation due to its high sensitivity, high throughput, less material-consumption, low cost, and enhanced spatio-temporal control.
Immunostaining
Immunostaining experiments are mult-istep protocols to detect specific antigens in biological samples. The sample is successively incubated or exposed to fixation agents, washing buffers, and probes.
In conventional protocols on petri dishes or well plates, fluid deliveries are performed manually using pipettes. Solutions are directly added to the sample.
Transferring protocols from a Petri dish to a microfluidic format usually requires some minor adjustments, as cells or tissue samples are exposed to solutions differently. Therefore, the use of microfluidic for cell biology provides many advantages by replacing the use of Petri dishes with microfluidic chambers (closed systems). Consequently, solutions cannot be deposited directly on the top of the cells but are perfused over the sample at a controlled flow rate, reaching all cells homogeneously. The use of a rotary valve with a perfusion system enables one to perfuse different solutions at given time points in the chip or chamber. In addition, automating the sequential delivery of solutions saves time and avoids creating bubbles inside the chips by disconnecting and reconnecting the traditional pump to the chip.
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How important is the microlfluidic flow control in cell biology?
In drug discovery, it‘s important to control the flow rate of the sample used. As multiple formulations are tested, experimenters must be able to control flow rate while keeping the same level control of fluid handling at each stage of the experiment. DNA and RNA hybridization are often performed under a microscope. Manual injection over a microscope can be risky for the following reasons:
- A touch of pipette cone can misplace the dish and misrecord the position
- Samples can be flushed away during pipetting
- Liquid can be spilled over the microscope
Flow Controllers Available on the Market
Syringe Pumps
Multiple flow control technologies are available for sub-millimeter range fluid management. Syringe pumps are commonly used for fluid control. However, in microfluidics, particularly in microfluidics for cell biology, the use of syringe pumps is often problematic because the flow is not constant. This discontinuity is due to the mechanical system that creates an oscillating flow.
Manual Injection
Current methods for multiple fluid delivery, where microscopy techniques are simultaneously required and mostly made by manual pipetting action include , Immunolabeling, and DNA and RNA hybridization.. Manual injection can lead to uncontrolled and non-homogeneous fluid velocity, which can damage samples.
| Pipette | Aria | |
|---|---|---|
| Type of injections | Abrupt injections (up to 1mL in few seconds) Disparate injections Turbulent flow | Smooth & controled injections Identical injections Laminar flow |
| Geometry at injecion tip | Conic shape: Important shear strain Unhomogeneous fluid velocity | Straight shape: No modification at the injection tip |
Fluigent Pressure-Based Flow Control
As the demand for microfluidic pumps with higher flow stability, fast response time, versatility, and automation capabilities has increased, pressure controllers have become the device of choice for many users.
The working principle of such pumps is to pressurize the sample reservoirs to control the pressure drop between the inlet and the outlet of the microfluidic system. The responsiveness of the generated flow rate depends on the responsiveness of the pressure pump.
To overcome these issues, Fluigent’s Aria automates multiple fluid deliveries. The sample is preserved as the flow rate is controlled. Reproducibility increases as inter and intra operator variability are eliminated.
