Putting Cells into Cryopreservation
If you work with cells, you know that one of the biggest challenges is keeping good viable stocks of your cells. One of the best ways to do that is through cryopreservation, which is the process of freezing cells so that they can be thawed and used later. But if you're going to freeze cells, you need to do it right - and that means using the right lab equipment.
Fear not, we're here with tips and tricks to keep your cells happy, healthy, and frozen.
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Why is Cryopreservation So Important?
Cryopreservation allows you to store cells for long periods without worrying about them dying. If you're working on a long-term project, want to store protein-expressing clones, or just need to keep cells around for future experiments, cryopreservation is the way to go.
But as with any technique, cryopreservation has its challenges. One of the biggest challenges is that freezing can cause damage to cells, which can result in decreased viability and altered cell function when they're thawed. This is why it's important to use the right labware and techniques.
What Labware Do I Need for Cryopreservation?
Cryovials are a must-have. They are small plastic or glass tubes designed to hold cells for freezing. You'll also need a cryoprotectant; a solution to help protect cells from damage during the freezing process. Common cryoprotectants include glycerol.
But other lab equipment and essentials can make cryopreservation easier and more effective. Here are a few:
Insulated Containers
Lab coolers are insulated containers that can hold cryovials and are designed to maintain the low temperatures necessary for sample storage, usually in the range of -80 to -196 °C. They come in a variety of sizes and styles, from small benchtop boxes that can hold just a few vials to larger tanks that can hold hundreds.
Pens and Markers
Permanent markers are used to label cryovials with important information such as cell type, passage number, and freezing date. It may seem like a small thing, but labeling your vials can make a big difference when it comes to locating them in the cold stores.
Cryogenic Apparel
Cryogenic apparel is designed to protect your skin and the cells you are working with during cryopreservation. Our cryogenic apparel consists of materials that can withstand low temperatures and protect against cryoprotectants.
Cryogenic Vials and Caps
Used for the storage of cryopreserved samples, cryogenic vials are made of materials that can withstand the low temperatures required for sample storage. They are usually made of polypropylene, which has a low-temperature resistance and is compatible with most cryoprotectants. Cryogenic vials come in different sizes, from small volumes of a few hundred microliters to larger volumes of several milliliters.
Labels
Labels are crucial in cryopreservation, especially when it comes to identifying samples. Cryogenic vials and containers should be labeled with permanent markers or labels that are resistant to low temperatures and can withstand cryogenic storage conditions.
Each label should contain essential information such as the date of collection, the type of sample, the passage number, and the identity of the researcher who collected the sample. A clear and legible label can prevent sample mix-ups and ensure accurate identification.
Frequently Asked Questions
Q:
How do I prepare cells for cryopreservation?
A:
Cells need to be prepared carefully before cryopreservation to improve survival. Ideally, cells should be in the logarithmic growth phase, usually subconfluent, and healthy. Detach the cells using an appropriate enzyme (typically trypsin) or mechanical method, resuspend them in a cryoprotective medium like dimethyl sulfoxide (DMSO), and then freeze them at a controlled rate. The cells should be completely free of serum to prevent the formation of ice crystals.
Q:
Can I freeze cells immediately after seeding?
A:
We would not recommend freezing cells immediately after seeding. Cells need time to adhere, spread, and establish a healthy monolayer before freezing. Waiting for a few passages or population doublings is advisable before considering freezing again. It’s important to try and freeze cells at the same stage each time to improve experimental reproducibility.
Q:
Is it possible to freeze primary cells?
A:
This will likely depend on the type of primary cell. While some primary cells can be cryopreserved, it can be more challenging compared to immortalized cell lines. Like any cell, it's important to consider the cell type, passage number, use of cryoprotectants, controlled cooling rates, and rapid thawing. Primary cells might show slower growth and altered behavior initially after thawing, but they can adapt over passages. If possible, consider testing the cryopreservation process by freezing cells at different densities, passage numbers and using different cryoprotectants.
How long will my cells be cryopreserved?
The length of time that cells can be cryopreserved depends on several factors, including the type of cell, the method of cryopreservation, and the conditions under which the cells are stored.
Generally, most cell types can be cryopreserved for long periods of time, often several years or even decades, if they are stored properly.
However, not all cells are equally amenable to cryopreservation, and some may lose viability or function after thawing. Additionally, the longer cells are cryopreserved, the greater the chance of damage to the cells, which can affect their viability and function.
Primary cells are particularly difficult to cryopreserve. But as with all cells, we recommend you test the viability and function of cells after cryopreservation and thawing, and periodically monitor the cells during long-term storage to ensure their quality is maintained.
Should I freeze my cells quickly or slowly?
A slower rate of freezing is generally preferred for most types of cells, because it reduces the likelihood of damage from ice crystal formation. Slow freezing with a cryoprotective agent allows the agent to penetrate the cells and prevent ice crystal formation while rapid freezing can cause damage and reduce viability.
