Criocutting
Freezing the tissues is a quick way to preserve and harden them without the need for inclusion.
It has the following advantages:
Molecular preservation is maximum.
Frozen samples can be cut into thin and thick sections
This is possibly the fastest way to obtain sections since it is possible to freeze the sample without fixation and cut it immediately, such as biopsies.
To properly preserve the structure of the tissue, it must be frozen very quickly, usually in isopentane cooled with liquid nitrogen, which prevents the formation of ice crystals that damage cellular structures.
The samples may also be fixed and then frozen after incubation in antifreeze solution. This prevents the formation of large ice crystals during freezing, which can damage the cell structures. This freezing allows even the cell ultra-structure to be preserved.
To preserve the structure and volume of the samples, they can be embedded in gelatine or glycol and resin-based compounds and then frozen, forming a block that facilitates the handling of the sample and also protects it and once the cut is obtained also facilitates its handling.
The cryostat is used to obtain freezing sections of variable thickness (5 to 30 µm). In this apparatus the entire cutting system (usually a rotating microtome) is enclosed in a refrigerated chamber whose temperature can be regulated below 0°C. It is advisable to place the sample in the cryostat chamber until the temperature of the sample is equal to that of the cryostat in order to obtain homogeneous sections. This allows the sample to be handled at low temperature and thus obtain the cuts without any variation in the process temperature.
The sections thus obtained are collected and processed for subsequent observation in accordance with the provisions of the protocol followed.
It has the following advantages:
Molecular preservation is maximum.
Frozen samples can be cut into thin and thick sections
This is possibly the fastest way to obtain sections since it is possible to freeze the sample without fixation and cut it immediately, such as biopsies.
To properly preserve the structure of the tissue, it must be frozen very quickly, usually in isopentane cooled with liquid nitrogen, which prevents the formation of ice crystals that damage cellular structures.
The samples may also be fixed and then frozen after incubation in antifreeze solution. This prevents the formation of large ice crystals during freezing, which can damage the cell structures. This freezing allows even the cell ultra-structure to be preserved.
To preserve the structure and volume of the samples, they can be embedded in gelatine or glycol and resin-based compounds and then frozen, forming a block that facilitates the handling of the sample and also protects it and once the cut is obtained also facilitates its handling.
The cryostat is used to obtain freezing sections of variable thickness (5 to 30 µm). In this apparatus the entire cutting system (usually a rotating microtome) is enclosed in a refrigerated chamber whose temperature can be regulated below 0°C. It is advisable to place the sample in the cryostat chamber until the temperature of the sample is equal to that of the cryostat in order to obtain homogeneous sections. This allows the sample to be handled at low temperature and thus obtain the cuts without any variation in the process temperature.
The sections thus obtained are collected and processed for subsequent observation in accordance with the provisions of the protocol followed.
Instrumentation
Staff
Luis Miguel Alonso Colmenar
Carmen Hernández López
Carlos Pérez Martín
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CAI Técnicas Biológicas
Flow Citometry and Fluorescence Microscopy Unit
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