Cryopreservation CSIR NET life sciences
Summary
TLDRThis tutorial video delves into cryopreservation, the process of preserving biological tissues at extremely low temperatures, typically -195°C, using liquid nitrogen. It highlights the importance of this method for halting cellular activity to prevent cell death, and discusses the challenges of avoiding ice crystal formation which can damage cells. The concept of vitrification is introduced as a gradual temperature increase process using vitrifying agents to safely thaw preserved tissues without cell rupture, ensuring successful tissue preservation.
Takeaways
- 🧊 Cryopreservation is the process of preserving biological specimens at very low temperatures, typically around -195°C, which is the boiling point of liquid nitrogen.
- 🌡 The extreme cold is necessary to halt all cellular processes, preventing any enzymatic activity that could lead to cell death.
- 🧬 Biological tissues, including tissues and organs, are preserved to maintain their viability for future use, ensuring that they remain 'alive' during storage.
- ❄️ Liquid nitrogen is commonly used in cryopreservation due to its extremely low temperature, which is crucial for stopping all biological activity within the tissues.
- 🔬 If tissues are stored at higher temperatures, cellular activities such as enzyme functions and metabolic processes would continue, leading to cell death and failing the purpose of preservation.
- 💧 The risk of ice crystal formation during cryopreservation can damage cells; rapid freezing helps to mitigate this by preventing water from expanding and rupturing cell membranes.
- 🔬 Vitrification is a process used to slowly transition tissues from cryopreservation temperatures to normal temperatures, using agents that prevent the formation of damaging ice crystals.
- 🌡 The vitrification process involves a gradual increase in temperature, avoiding rapid changes that could cause cell membrane rupture and cell death.
- 🛠 Vitrifying agents are used to facilitate the controlled warming of tissues, ensuring stability and preventing damage during the thawing process.
- ♻️ The script emphasizes the importance of carefully managing temperature changes during both the freezing and thawing processes to maintain the integrity of biological samples.
- 📚 The tutorial aims to provide a basic understanding of cryopreservation, highlighting the significance of temperature control and the role of vitrification in preserving biological tissues.
Q & A
What is cryopreservation?
-Cryopreservation is the process of preserving biological specimens or samples, such as tissues, organs, or cell lines, at extremely low temperatures, typically around -195°C, to maintain their viability for future use.
Why is liquid nitrogen commonly used in cryopreservation?
-Liquid nitrogen is used because its boiling point is -195.8°C, which provides the ultra-low temperatures necessary to halt all cellular processes and prevent cell death, making it ideal for preserving the viability of biological tissues.
What is the significance of storing biological tissues at -195°C?
-Storing at -195°C halts all cellular processes, including those that could lead to cell death such as apoptosis. This temperature is crucial to prevent the activation of enzymes that could degrade the tissue or lead to cell death.
What is the risk associated with rapid freezing of biological samples?
-Rapid freezing can lead to the formation of ice crystals within the tissue. The expansion of water as it freezes can cause cell membranes to rupture, leading to cell death, which is why a controlled freezing process is essential.
What is vitrification in the context of cryopreservation?
-Vitrification is the process of slowly warming a cryopreserved tissue from ultra-low temperatures to normal temperatures using vitrifying agents. This gradual increase in temperature helps prevent damage to the tissue caused by rapid temperature changes.
What role do vitrifying agents play in the warming process of cryopreserved tissues?
-Vitrifying agents are used to prevent the formation of ice crystals during the warming process. They help to stabilize the tissue and allow for a controlled, gradual increase in temperature, reducing the risk of cell damage or death.
Why is it important to avoid rapid temperature changes when thawing cryopreserved tissues?
-Rapid temperature changes can cause significant stress to the cells, potentially leading to cell membrane rupture and cell death. A controlled, gradual increase in temperature helps to minimize these risks.
What happens to cellular activities at temperatures above -195°C?
-At temperatures above -195°C, cellular activities such as enzyme functions and metabolic processes can begin to occur, which could lead to cell death if the cells are not intended to be active.
How does the process of vitrification help maintain cell stability during thawing?
-Vitrification involves a step-by-step increment of temperature, which allows the cells to gradually adjust to the change, thereby maintaining cell stability and reducing the risk of damage.
What is the typical temperature range for the vitrification process?
-The vitrification process involves gradually increasing the temperature from -195°C, possibly through stages such as -50°C, -25°C, 0°C, and finally to 35°C, depending on the specific requirements of the tissue being thawed.
Why is the preservation of living tissues important in cryopreservation?
-The preservation of living tissues is important to ensure that the biological specimens remain viable for future use, such as in transplantation or research, without the risk of cell death or degradation.
Outlines
🧊 Cryopreservation Basics and Temperature Significance
This paragraph introduces the concept of cryopreservation, which is the preservation of biological samples at extremely low temperatures. The primary focus is on the significance of using temperatures as low as -195°C, which is the boiling point of liquid nitrogen. The purpose of such low temperatures is to halt all cellular processes, preventing cell death due to apoptosis or other enzymatic activities that could occur at higher temperatures. The paragraph also touches on the importance of rapid freezing to prevent the formation of water crystals that could damage cells.
🔄 The Vitrification Process in Cryopreservation
The second paragraph delves into the process of vitrification, which is essential for safely thawing cryopreserved biological samples. It explains the need for a gradual temperature increase to avoid cell damage that could occur from rapid temperature changes. Vitrification agents are used to facilitate this slow and controlled temperature rise, ensuring the stability of the cells. The paragraph outlines the step-by-step process of incrementally raising the temperature from -195°C to physiological levels, emphasizing the avoidance of a direct and rapid transition that could be detrimental to cell integrity.
Mindmap
Keywords
💡Cryopreservation
💡Liquid Nitrogen
💡Cellular Activities
💡Apoptosis
💡Vitrification
💡Vitrifying Agents
💡Temperature Control
💡Enzymes
💡Cell Membrane
💡Biological Tissues
Highlights
Cryopreservation is the process of preserving biological specimens at extremely low temperatures.
The term 'cryopreservation' originates from 'cryo' indicating cold and 'preservation' for maintaining the state of the samples.
Biological tissues, organs, and cell lines can be preserved through cryopreservation.
The ideal temperature for cryopreservation is approximately -195°C, which is the boiling point of liquid nitrogen.
Liquid nitrogen is commonly used in cryopreservation due to its extremely low temperature.
At -195°C, cellular activities and enzymatic reactions are halted, preventing cell death.
Cryopreservation inhibits apoptosis or programmed cell death, thus preserving the viability of the tissue.
Vitrification is a process in cryopreservation that prevents the formation of water crystals which can damage cells.
Rapid freezing is crucial to prevent water from expanding and bursting the cell membranes.
The thawing process must be gradual to avoid cell membrane rupture due to rapid temperature changes.
Vitrifying agents are used to facilitate the controlled increase in temperature during the thawing process.
Incremental temperature increase helps maintain cell stability and prevent cell death post-thawing.
Cryopreservation is vital for the long-term storage of biological samples without loss of viability.
The process of cryopreservation halts all cellular processes, ensuring the preservation of the sample's integrity.
Cryopreservation has significant practical applications in medical and biological research.
Understanding the principles of cryopreservation is essential for successful preservation and revival of biological samples.
The tutorial provides a basic understanding of cryopreservation techniques and their importance in preserving biological tissues.
Transcripts
[Music]
welcome back friends in this video
tutorial we'll be talking about
cryopreservation it's just basic small
video about
cryopreservation
cryo
preservation okay so what do we mean by
cryopreservation as the term suggest
cryo preservation means preserving
biological specimens or samples like
tissues or different organs or different
cell lines in cold temperature because
when cryo is associated we always talk
about cold climate and cold temperature
so
preservation so
preservation
of tissue which is living
obviously in cold temperature now the
question comes how much cold right and
the answer is here it is almost let's
say - 195° c or let's say 77 Kelvin or
something so - 197 which is
below
below
Subzero temperature minus 195° cus now
why this particularly minus 195° C
because this is the boiling point
of of liquid
nitrogen boiling point of liquid
nitrogen okay so usually in cry
preservation of tissue grafts and C
preservation of biological tissue
specimens we usually use this liquid
nitrogen for that okay now liquid
nitrogen is very cold because you can
see minus 195° C very very very cold uh
in temperature now why we require that
much very small amount of uh temperature
to store them we can store it freezing
temperature like 0 -4 -5 - 10 but is -
195° c and the answer for that the key
for the cry preservation is that we need
to store the biological tissues which
are living tissues so that they still
leave right because if you store them in
higher temperature the activity of there
all the cellular activities like
activity of enzymes like polymerous
enzyme for the replication transcription
different enzymes for metabolism they
will start to work because for all the
biological enzymes to work it requires a
particular temperature more than 0°
celus usually it acts better at
physiological temperature like 35 to 37°
CSUS many of them works well in the home
temperature like 25 to 30° C but very
few of them can also Act on that 5 10
12° cus temperature so if we store them
in low temperature like 510 or close to
zero it will still act on it and the
cell will finally go and die right
because for the cell death it requires
some enzymes some deadly Pathways called
apoptosis or programmed cell death right
so if we put if those enzymes
responsible for program cell death are
activated at a particular temperature
and if you store this cell in that
temperature if those enzymes are active
then the cell will die and the purpose
for preserving tissue will be failed so
for that reason we have seen that it
atus 195° C temperature none of those
enzymes are active to kill the cell or
to guide the cell through apoptosis so
all of the processes I repeat all of the
cellular processes that are usually
going on inside the cell are kind of
halted are kind of in inhibited at that
low temperature that you have talked
that's why we used to store our uh
samples in that much cold temperature
okay that's the reason and second thing
another question CSI net people can ask
you is that what is vitrification now
let me explain this term
vitrification now one important thing
about this C preservation in any kind of
cold preservation is once you put it
into that high uh very low temperature
it there's a chance of water crystal
formation in your tissue now if water
crystal form we know that at Crystal
form at at the solid form at the icy
form water expands more so it can burst
your cell but usually for that reason we
need to rapid rapid freeze them in this
temperature that's a very important
point now let's say you've made the
sample you've taken the sample prepare
the graph and you put it rapidly into
the freezing Machinery everything is
okay everything is fine it is stored it
is preserved now you need to take that
particular sample out from that cold
temperature and you need to PL place it
in the normal temperature right so what
you need to do we need to increase the
temperature now remind you in any
condition if you increase the
temperature rapidly just like freezing
it rapidly it will kill the cell why
because increasing temperature rapidly
will cause certain changes very fast
into your cell and the cell membrane
will not be able to hold on to all those
changes at very fast so the cell
membrane will rupture and the cell will
die to prevent that what we need to do
we need to increase the temperature
little by little with time right and
using certain amount of different agents
different watering agents different uh
water supplying agents and different
enzymatic agents now those agents are
called vitrifying agents and the process
of slowly taking out of the tissue graph
from the liquid nitrogen to the normal
temperature is called the vitrification
process in this process what we do we do
stepbystep increment of
temperature so we take let's say it's
previously 195° cius temperature we add
some vitrification agent or vitrifying
solution one for example and the
temperature comes like let's say minus
50 then slightly let's 25 then it's kind
of zero then from it slightly we're
increasing towards 35° C and this is the
process not directly we're not going
this this path because this pathway is
very very dangerous this pathway is
dangerous so it is not taken either we
take the small stages because it helps
uh to maintain the St cell stable okay
so that is the vitrification process
okay so that's kind of it I hope that's
helpful thank you thank
you
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