Certain eukaryotic animal cells, such as primary T cells, nerve cells, and stem cells, have properties that make it difficult to obtain acceptable transfection effects by chemical methods such as the widespread use of lipofections. The cumbersome procedures and potential hazards of virus transfection make large-scale application of viruses to solve the transfection experiments of these cells become a path that experimenters are not willing to choose.
15 years ago, Amaxa technology appeared, improved the ancient electro-rotation technology, made electric shock waveforms and energy optimization for eukaryotic cells, and specially proposed a protective liquid (transfection liquid) formula for protecting cells at the moment of electric shock, thereby greatly improving these The efficiency and survival rate of primary eukaryotic cell electroporation, the DNA entering the cell is extremely fast due to electroporation, and protein expression can also be observed within a few hours, thus amaxa company specifically "made" a word "nuclear transfection" "To illustrate the advantages of this technology. Thus, Amaxa defines the second generation of electrical technology. (Amaxa was later acquired by Lonza and gradually became known as the Lonza nuclear transfection instrument)
Amaxa technology was a revolutionary improvement 15 years ago, but with the increasing use of amaxa electrical equipment, it has gradually exposed some of its inherent flaws.
1) Amaxa technical support often says: "This is a problem with your cells."
The amaxa technology is based on the so-called "standard" cells of the US ATCC, but in fact the cells used in the laboratory are passed down from generation to generation by the laboratory or friendly laboratory, and they have been different from the ATCC standard cells. For the electric rotation, dancing on the tip of the knife: "increasing one point, the cell is electrocautery, reducing one point, the cell power does not go in." The nuance of the cell trait leads to a huge difference in the electrotransfer effect, which is why Most people don't get the electrical conversion effect on the amaxa standard experiment protocol: others are obviously 60% transfection efficiency. Why do I only have 30% of my life?
Because the Amaxa device encodes the transfection program according to the cell type, and in order to prevent the technology from leaking, it adopts the measure of over-protection. Each program code is completely garbled and alphanumeric, which makes it impossible for the user to perform the amaxa program for the laboratory cells. Optimize the adjustment. That is to say, once the transfection effect is not satisfactory, and other factors are excluded, if the cells are confirmed to be non-standard, Amaxa/Lonza technicians can only suggest that you buy ATCC standard cells to meet the effect of achieving amaxa technology. This is a typical “cutting footâ€, and the opinions of global experimenters are very large.
2) considerable cell death
Amaxa technology takes into account the problems of electric shock and chemical protection reagents, but is still conservative in the design of the most basic electric shock cups, using the oldest slit U-shaped electric shock cup (the ordinary electric shock cup used in the first generation BioRad electric shock technology). This electric shock cup design will inevitably have the problem of a large number of cell electrocution in the liquid surface and the adherent area from the principle of electrical physics. Since the electrical and physical properties of the electric shock cup are the core of the electrical design of the entire instrument, the improved electric shock cup design will result in the redesign of the entire machine system, and the Amaxa equipment cannot be changed. More importantly, other electric shock cup designs have been patented by other companies. This includes the core design of the new domestic third-generation electric equipment mentioned below. The amaxa technology has been blocked, and the space for further improvement is blocked. .
3) The amount of transfected cells is too small
The Amaxa technology has only 100ul and a smaller 20ul system. The 100 ul system is also a maximum of 10 ^ 6 grades, which is not the same as liposome/virus transfection can adapt to 10^6-10^9. This is one of the important reasons why many people do not choose the path of the electric circuit. Since Amaxa has chosen the route of the slit U-shaped electric shock cup, this has caused it to fail to increase the amount of cell transfection. In the past two years, Amaxa/Lonza has introduced a new machine that can do 10^7-10^8 transfection at one time, but the basic principle is still based on the 100ul electro-transformation reaction system. It is nothing more than a complex fluid system. In this way, a 1ml or 10ml transfection that could be done once is divided into 10 consecutive or 100 transfections. It can be seen that this kind of wrong technical route leads to the complicated and expensive machine construction of this solution, and the cost of consumables is even more expensive: one or two thousand RMB consumables to do a transfection of 10^8 cells, can you accept it?
Now, in the high-tech field of cell biology in this subdivision of electric power, the scientific research power of the Chinese has emerged! It has been commercialized. Has joined the fierce market competition and is facing the challenge of amaxa technology! It has been verified in many important research institutes in the United States and China (both laboratories with multiple amaxa equipment and many years of experience in amaxa use), successfully rewriting the views of users and achieving the victory in the commercial market competition.
 | Dr. Chen Jian, CEO, inventor of Celertrix electro-transfer technology. Graduated from Tsinghua University with a bachelor's degree in biology; received a Ph.D. in the Department of Pharmacology at Cornell University/Sloan-Kettering Cancer Center in 2004; and then worked on a screening model for new drugs for blood and immune diseases at the University of California, San Diego, Mount Sinai School of Medicine Engaged in the study of genetic recombination of immune cells. Received NIH funding to develop new electro-transfer technology and achieve breakthrough success |
Celetrix electric converter, there are several core technological breakthroughs.
First, the most important core innovation: the replacement of the traditional U-shaped electric shock cup design with a pressurized electric shock tube.
Adding a piezoelectric tube looks like a lot of cryotubes used in biological experiments, but turning it into a shock tube has never been thought of. This unique innovation, which draws on both the capillary electric field and the closed electrochemical effect, makes the transient of the eukaryotic cells shocked extremely stable and reliable, and is the fundamental principle of innovation. This has laid the foundation for its superior principle of higher electrical turn efficiency and higher cell survival. From the practice point of view, the pressurized electric shock tube rarely observes the "white foam" of cell death after the instantaneous large voltage produces the electric rotation effect, which fully demonstrates the correctness of its core theory, and further improves and develops this new era. Electrical technology provides the basic physical foundation. At present, this electric shock tube design has applied for US and Chinese patents, thus blocking the space for further improvement of amaxa technology.
Second, large-capacity cell electrotransformation can be achieved by simply expanding the volume of the pressurized electric shock tube.
This new generation of design of the electric shock tube provides a possibility that, as long as the control ability of the large energy current is obtained, it is theoretically possible to realize a one-time large-capacity cell electrotransfer by simply increasing the volume of the electric shock tube. At present, Celetrik has been able to provide 1ml (about 10^8 cells) of one-time shock, regardless of the reliability of the results (one turn, not 10 times 100ul), easy to use, and greatly reduce the cost of consumables. This is the inevitable development trend of large-capacity cell power transfer in the future. And as long as the circuit continues to improve, you can get a one-time turn of a larger number of cells, let us look forward to the launch of new Celetrix equipment.
Third, for the defects of Amaxa technology, the user's self-adjusting electrical parameters are called easy to use.
By simply adjusting one or two values, the user can fine-tune the cell electroporation conditions to obtain the optimal "electrical cell of their own laboratory", rather than this type (eg, the immobilization procedure for human T cells on Amaxa devices). , the parameters of the fixed electrical parameters, so that the best results can be obtained in accordance with the current laboratory cell conditions.
Fourth, for the very high cost of reagent supplies for Amaxa technology, Celertrix has significantly reduced the cost of daily use, only about 50-60% of the Amaxa equivalent (100ul system). If the user transfers more capacity, Celertrix provides 200ul once, and 1ml once capacity, then the savings of reagent supplies is only a fraction or even a tenth. This is the cost advantage of core technology improvements.
At present, Celetrim has just interviewed for less than two years, but there are already Yale University, Harvard University, NIH; there are also nearly 100 laboratories in China's domestic Chinese Academy of Sciences, Peking University, etc. These users are heavily used by the original Amaxa technology. They understand the difference between the two. Their choice is not only the trust of Chinese technology and products, but also the recognition of the superiority of the third-generation electric technology (if Amaxa technology is the second-generation electric technology).
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