Are you experimenting with chemical solutions in cell experiments?
Have you ever thought about:
Whether or not to mix after the chemical solution administration?
What is the difference in the first place in the first place?
In this article, I will explain how I administered the chemical solution in the dish and verified the situation of the chemical solution, both without mixing and by mixing. Please feel free to refer to my research, by any means, or likewise get in touch…
Table of contents
What is a drug test?
The role of DMSO
The need for mixing
Mixing during the observation stage
Additional benefits of automation
1．What is a drug test?
In this context, it is a test to see the reaction by administering the drug to the cells in the culture. The types of drugs include those that confirm the efficacy of drug candidates; substrates that illuminate cells and stains.
The addition of drug candidates is executed when performing drug efficacy and pharmacological tests on target cells. The main evaluation seems to be the determination of the life and death of the cell (i.e. how long it takes for the cell to die).
The addition of substrates and staining solutions that illuminate cells is performed when you want to analyze the state of the target cell in greater detail. Evaluation is often carried out by fluorescence microscopy.
2．The role of DMSO
In general, to add the above drugs to cells in culture, either a small amount is added directly to the culture supernatant, or the culture supernatant is replaced with a drug-containing one. Dimethyl sulfoxide (DMSO) is often used as a solvent when adding small amounts directly.
DMSO, which is familiar to people who have cultured cells and added chemical solutions during the process, is an amphiphilic, meaning it dissolves in water and oil, so it is always used when dissolving fat-soluble drugs that are difficult to dissolve in water and in the medium.
In fact, we verified the addition of oily ink that resembles a fat-soluble drug to water that resembles a medium (see video below).
Oily ink mixed with DMSO dissolves well in water when pipetting and becomes uniform, but only the oily ink separates (i.e. it does not mix).
DMSO is amazing! I didn't think it would be so different.
3．The need for mixing
When replacing the culture supernatant with a drug, there is no problem because the drug is mixed evenly in advance, but when adding a small amount directly to the culture supernatant, it is necessary to mix them, for the drug to spread evenly throughout.
That's where the pipetting is often done.
How much pipetting is actually mixed?
I verified this using oily ink that resembled a drug. (Video below)
It may seem obvious, but it mixes well when pipetting. On the other hand, a dish without pipetting may seem to be spread by diffusion at first glance, but if you look at it from the perspective shown at the end of the video, you can see that there is a considerable difference in concentration between the top and bottom.
It therefore seems that mixing. when adding a small amount, is essential.
4． Mixing during the observation
When administering the drug in small quantities, it was found that mixing (pipetting) was essential.
So, can this mixing operation be done in real-world experiments?
Most experiments with microscopic observation assume the following workflow:
Open the lid, on the microscope stage
These series of tasks must be performed with a high degree of accuracy, without moving the cells being observed.
If you add the drug to the cells in the culture and perform the same work, you will be required to operate very carefully.
If it was a more valuable sample, my hands would be trembling even more!
TOKAI HIT’S cellular perfusion system allows you to do this task and mix without touching the sample.
Here's what I actually tried:
As with manual pipetting, you can see how the ink resembles a drug that is evenly mixed.
At the end of the video, the up-and-down uniformity is also maintained.
In this case, it seems that it isn't necessary to operate administration and mix, while suppressing trembling hands (!).
5．Additional benefits of automation
In addition to being able to mix administration without touching the sample, the cell perfusion system can perform programmable operations, allowing imaging immediately after administration (0 seconds).
It is very useful for when you want to observe the rapid reaction inside the cell.
In addition, the program can be operated by exchanging signals with microscopes and other peripheral devices (i.e. receive and operate a signal and pass the signal after it operates).
I will elaborate on this separately, in a different article.
To summarize the above,
DMSO used for chemical solution administration is amazing at dissolving fat-soluble chemical solution in a medium.
Mixing is essential after chemical administration
Administration and mixing during observation of the microscope is difficult to perform manually
I trust that all makes sense!?
I hope this information will be useful to those who are doing some kind of chemical solution administration experiment to cells.
We look forward to your request.
We propose the optimal perfusion environment by combining lineups and technologies.
Thank you very much.