Related Experiment Video

Updated: Jan 25, 2026

Nanosponge Tunability in Size and Crosslinking Density
11:15

Nanosponge Tunability in Size and Crosslinking Density

Published on: August 4, 2017

8.1K

The impact of crosslinking and non-crosslinking fixatives on antigen retrieval and immunohistochemistry

Cornelia Stumptner1, Daniela Pabst1, Martina Loibner1

  • 1Diagnostic- and Research Center for Molecular Biomedicine, Institute of Pathology, Medical University of Graz, Graz, Austria.

New Biotechnology
|May 15, 2019
PubMed

Pre-analytical factors can greatly influence the outcome of molecular analyses in medical diagnostics and research. This also applies to in situ staining techniques such as immunohistochemistry (IHC), where different types of tissue fixation methods lead to different modifications of proteins and thus can affect differently the detection by antibodies. For formalin-fixed paraffin-embedded (FFPE) tissue, antigen retrieval is applied in order to reverse the negative effects of formalin and re-establish immunoreactivity. Most antibodies and protocols used in IHC are optimized for FFPE tissue, but not for paraffin-embedded tissue treated with other fixatives such as non-crosslinking fixatives. We report results from systematic studies on distinct pre-analytical conditions in IHC, immunofluorescence and electron microscopy. Parameters investigated are the impact of crosslinking and non-crosslinking fixatives (comparing formalin and PAXgene Tissue fixation) on whole tissue, subcellular structures and organelles, as well as on ultrastructure. The results generated show that minor changes in antigen retrieval conditions may have a major impact on IHC results and that protocols optimized for crosslinking fixatives may not be used for other fixatives without re-validation. Key antigen retrieval parameters such as buffers with different pH and duration of microwave treatment must be tested systematically for each antibody and fixation protocol.

Keywords:
Antigen retrievalFormalin-fixed paraffin embedded tissueImmunohistochemistryIn situ detectionPAXgeneStandardization

More Related Videos

Preparation of DNA-crosslinked Polyacrylamide Hydrogels09:06

Preparation of DNA-crosslinked Polyacrylamide Hydrogels

Published on: August 27, 2014

15.3K
Author Spotlight: Quantitative Detection of DNA Protein Crosslinks and Their Post-Translational Modifications10:12

Author Spotlight: Quantitative Detection of DNA Protein Crosslinks and Their Post-Translational Modifications

Published on: April 21, 2023

3.6K

Related Concept Videos

Retrieval01:12

Retrieval

Retrieval is the process of getting information out of memory storage and back into conscious awareness. This ability is essential for daily tasks like brushing hair and teeth, driving to work, and performing job duties. Retrieval occurs in three ways: recall, recognition, and relearning.
Recall involves accessing information without cues, such as during an essay test, where individuals must retrieve facts and concepts from memory unaided. Another example is remembering the name of a colleague...
425
ER Retrieval Pathway01:45

ER Retrieval Pathway

In the secretory pathway, vesicles transport proteins from one cellular compartment to another in forward transport to deliver the protein to its correct location. Occasionally, misfolded proteins and incorrect proteins escape their original compartments, and a retrieval pathway is used to return the escaped proteins to their original compartment.
The ER uses many checkpoints to prevent the entry of incorrectly folded or a resident protein as cargo onto a transport vesicle. These mechanisms...
4.7K
Fixation and Sectioning01:03

Fixation and Sectioning

Two basic types of preparation are used to visualize specimens with a light microscope: wet mounts and fixed specimens.
The simplest type of preparation is the wet mount, in which the specimen is placed in a drop of liquid on the slide. A liquid specimen can be directly deposited on the slide using a dropper. Solid specimens, such as skin scraping, can be placed on the slide before adding a drop of liquid to prepare the wet mount. Sometimes the liquid is simply water, but stains are often added...
7.5K
Impact of Groups on Groups01:19

Impact of Groups on Groups

Social psychologists analyze how groups influence one another, shaping social structures and interactions through both cooperation and competition. These dynamics manifest in various ways, ranging from economic partnerships to intergroup conflicts that shape societal structures and perceptions.Cooperation and Competition in Intergroup RelationsIntergroup relationships vary across contexts, sometimes fostering cooperation and mutual benefit while at other times leading to conflict and...
237
Immunocytochemistry and Immunohistochemistry01:22

Immunocytochemistry and Immunohistochemistry

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.
These...
13.6K
Carbon-dioxide Fixation01:28

Carbon-dioxide Fixation

Carbon dioxide fixation in prokaryotes enables the assimilation of inorganic carbon into organic molecules, supporting biosynthetic pathways, sustaining ecosystems, and contributing to the global carbon cycle. It also has industrial applications in carbon capture and bioproduct synthesis. Autotrophic organisms rely on this process to utilize CO₂ as a carbon source in diverse environments.The Calvin CycleThe Calvin cycle is the most widespread carbon fixation mechanism, primarily used by...
682