Cell biology & Microscopy
The advantages of flow control for Cell biology and Microscopy
- Shear stress control
- Automation of multiple fluid delivery
- Long term perfusion
Main applications
RNA and DNA hybridization
Over the past 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…).
Recently, 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 really paved the way for this approach to these new therapies due to the failure of a ‘one size fits all’ 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 drug combination.
Immunostaining
Immunostaining experiments are multistep 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 pipets. Solutions are directly added to the sample.
Transposing protocols from a petri dish to a microfluidic format usually requires some minor adjustments as cells or tissue samples are exposed to solutions in a different manner. As opposed to petri dishes, microfluidic chips or chambers are closed systems. Therefore 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 homogenously. The use of a rotary valve with perfusion systems 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.
Resources
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Application notes Capture and Labeling of cancer cells using Aria Read more
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Application notes Peristaltic pump vs pressure-based microfluidic flow control systems for Organ on-chip applications Read more
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Expertise reviews Comparison between peristaltic, syringe and pressure pumps for microfluidic applications Read more
Cell biology and flow control
In drug discovery it‘s important to control the flow rate of the sample used. As multiple formulation are tested it is important to have the possibility to control at the same time the flow rate while keeping the same level control of fluid handling at each stage of experiment. DNA and RNA hybridization are often performed under a microscope. Manual injection over a microscope can be risky as :
- a touch of pipette cone can misplace the dish and recorded 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, 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. This can be problematic for certain experiments such as biological ones.
Manual injection
Current method for multiple fluid delivery where microscopy technics are require at the same time are made mostly by manual pipetting action (Immunolabelling, DNA and RNA hybridization… ). Manual injection can lead to uncontrolled and non-homogeneous fluid velocity which can damage your sample.
Fluigent pressure-based flow control
As demand for microfluidic pumps with higher flow stability, fast response time, versatility, and automation capabilities have increased, pressure controllers have become the device of choice
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, Aria automates multiple fluids delivery. The sample is preserved as the flow rate is controlled. Reproducibility is increased as inter and intra operator variability are eliminated.
