Fluorescent CRISPR-Cas9 complexes injected into mouse primary hepatocytes with the CRISPR system.
Genome engineering by the CRISPR-Cas system is traditionally carried out by delivering plasmids encoding for the CRISPR components by lipofection or electroporation to the cytoplasm of the targeted cells. This approach, although successful in easy to transfect cells, has two mayor disadvantages: 1) The plasmid DNA has to be translated to proteins and delivered to the nucleus. 2) Electroporation and lipofection is cytotoxic and can lead to excessive cell death.
Nucleofection of pre-assembled Ribonuclear-Protein complexes (containing Cas proteins, guide RNA and optionally a DNA-repair template) overcomes the first disadvantage, but often leads to extensive cell death. This is especially critical when working with rare or hard-to-transfect cells such as iPSCs, neurons or cardiomyocytes.
With the FluidFM CRISPR system, deliver your CRISPR-Cas complexes directly to where they are required: the nucleus. This avoids the risk of degradation of the repair template in the cytoplasm, therefore increasing the chance of HDR event to occur.
Using a fluorescent tracer, you can ensure that 100% of your targeted cells will be delivered with your CRISPR-Cas complexes. Moreover, the gentle process of injection with the CRISPR system preserves cell viability over 95%.
Whether you are working with hard-to-transfect cells like iPSCs or a standard cell line such as CHO cells, you can selectively choose the cells you want to inject within a cell culture. The neighboring cells won’t be affected.
No toxic compounds, high viability (> 95%)
Targeted intracellular delivery
Directly inject selectively into nucleus or cytoplasm
Inject into hard-to-transfect cells
Fast & easy
Process hundreds of cells/hour with intuitive software
Boost CRISPR efficiency by direct delivery into the nucleus.
Get in touch with our experts and find out how our technology can help your research.
Traditional methods deliver the CRISPR-Cas components (Cas9 protein, guide RNA, DNA repair template) to the cytoplasm. However, as these components are unstable in this compartment and have to be transported to the nucleus, editing of the targeted locus becomes inefficient. With the CRISPR system, pass easily over the cytoplasmic and nuclear membranes in directly delivering the material to the nucleus, making sure that your CRISPR-Cas complexes and repair template reach their targets. This increases the chance of HDR events to happen.
Adherent cells can easily be isolated using a FluidFM micropipette. This is particularly relevant to analyze the success of your gene editing, or to create a monoclonal cell line from the successfully edited cells. Discover more about FluidFM cell isolation here.
With the integrated Volume Calculator feature, easily measure the number of complexes that you injected into a specific cell. This is particularly useful when you want to optimize the dosage to maximize on target and minimize off target effects.
Deliver as many gRNA as you want in a single nucleus, in a single injection.
Only 1µl of CRISPR-Cas solution is required to inject thousands of cells.
Our CRISPR system has already been successfully used to inject into various cell types such as iPSC, mouse or human ESC or primary hepatocytes. Find the full list here.
Process hundreds of cells/hour.
The tiny aperture of the FluidFM nanosyringe (600 nm), its smoothly controlled movement and the fact that no toxic compounds are necessary to allow the genetic material to enter the cell lead to a cell viability over 95%
Our software records the XY-coordinates of the injected cells and allows the user long-term observation of the modified cells with the desired settings (fluorescence filter, exposure time etc.).
HOW IT WORKS.
Through the microfluidic channel inside a FluidFM probe, CRISPR-Cas complexes can be dispensed through a sub-micrometer aperture at the tip of a cantilever. The sensitive force feedback system provides a reliable distinction between gentle contact with cell membranes and perforation, making it possible to inject directly into single cells and even selectively choose the nucleus or cytoplasm to be injected. Check here the video explaining the working principle of FluidFM nano-injection.
The specific probe used for the direct delivery of CRISPR-Cas complexes into a cell is our FluidFM nanosyringe. It enables injection into single cells without compromising cell viability. The very sharp apex and the around 600 nm aperture at the front side of the pyramidal tip enable gentle injection and allow to work with compounds with several molecular weights and densities.
For more information on the FluidFM technology, please see here.
FluidFM is a highly versatile tool with huge potential in the field of single-cell studies. The instrument enables us to address several unanswered questions in the epigenetics field by pushing experimental boundaries.