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Related Concept Videos

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Immunocytochemistry (ICC) and immunohistochemistry (IHC) are techniques that use antibodies to check for specific proteins or antigens in a sample. The technique was first published by Albert Coons in 1941 to detect the presence of pneumococcal antigen in tissue sections from mice infected with Pneumococcus. Immunocytochemistry helps localization of proteins or antigens in individual cells like blood cells, stem cells, etc., while immunohistochemistry does the same for tissue samples.
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Heating a crystalline solid increases the average energy of its atoms, molecules, or ions, and the solid gets hotter. At some point, the added energy becomes large enough to partially overcome the forces holding the molecules or ions of the solid in their fixed positions, and the solid begins the process of transitioning to the liquid state or melting. At this point, the temperature of the solid stops rising, despite the continual input of heat, and it remains constant until all of the solid is...
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When a liquid vaporizes in a closed container, gas molecules cannot escape. As these gas phase molecules move randomly about, they will occasionally collide with the surface of the condensed phase, and in some cases, these collisions will result in the molecules re-entering the condensed phase. The change from the gas phase to the liquid is called condensation. When the rate of condensation becomes equal to the rate of vaporization, neither the amount of the liquid nor the amount of the vapor...
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Gas pressure is caused by force exerted by gas molecules colliding with the surfaces of objects. Although the force of each collision is very small, any surface of an appreciable area experiences a large number of collisions in a short time, which can result in high pressure.
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Calorimetry is a technique used to measure the amount of heat involved in a chemical or physical process or to measure the heat transferred to or from a substance. The heat is exchanged with a calibrated and insulated device called the calorimeter. Calorimetry experiments are based on the assumption that there is no heat exchange between the insulated calorimeter and the external environment. The well-insulated calorimeters prevent the transfer of heat between the calorimeter and its external...
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Related Experiment Video

Updated: Jan 27, 2026

Biological Sample Preparation by High-pressure Freezing, Microwave-assisted Contrast Enhancement, and Minimal Resin Embedding for Volume Imaging
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High-pressure freezing for immunocytochemistry

P Monaghan1, N Perusinghe, M Müller

  • 1Institute of Cancer Research, Sutton, Surrey, UK. monaghan@icr.ac.uk

Journal of Microscopy
|January 29, 1999
PubMed
Summary

High-pressure freezing and slow acetone freeze-substitution offer superior preservation for ultrastructural immunocytochemistry. This method optimizes tissue architecture and cellular morphology for detailed cellular analysis.

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Area of Science:

  • Cell Biology
  • Microscopy Techniques

Background:

  • Ultrastructural immunocytochemistry demands minimal antigen damage during sample processing.
  • Conventional immersion fixation can harm antigens; rapid freezing offers advantages.
  • Ambient pressure rapid freezing limits sample thickness to ~15 µm, while high-pressure freezing allows up to 200 µm without ice crystal artifacts.

Purpose of the Study:

  • To evaluate different freeze-substitution protocols for high-pressure frozen samples in immunocytochemistry.
  • To optimize tissue architecture, resin embedding, and immunolabeling.
  • To determine the best post-freezing preparation for high-pressure frozen samples.

Main Methods:

  • High-pressure freezing of biological samples.
  • Comparison of various freeze-substitution media (acetone, methanol, tetrahydrofuran) and conditions (time, temperature).
  • Evaluation of tissue morphology, ease of embedding, and immunocytochemical labeling efficiency.

Main Results:

  • Freeze-substitution in acetone with 2% osmium tetroxide followed by room temperature epoxy-resin embedding yielded optimal morphology.
  • Slow warm-up and extended substitution times in acetone improved freezing quality and cellular preservation.
  • While methanol and tetrahydrofuran offered faster substitution, they resulted in poorer cellular morphology compared to acetone.

Conclusions:

  • High-pressure freezing combined with slow freeze-substitution in acetone provides excellent preservation for ultrastructural immunocytochemistry.
  • Optimized protocols enhance tissue architecture and cellular detail, crucial for accurate antigen localization.
  • Careful sample handling and slow warming are key to minimizing ice crystal damage.