Tissue fixation

For immunocytochemical localization of certain amino acid transmitter enzymes in the central nervous system, a good preservation of tissue morphology and antigen antigenicity is essential In general, perfusion fixation is better than immersion fixation. For perfusion fixahon one may perfuse the fixative into the animal circulatory system through the left ventricle of the heart or through the abdominal aorta. 

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Microwaves have successfully been used for rewarming of blood for medical appHcations (157). Another successful appHcation, not commetciali2ed as of this writing, is the use of microwave heating for rapid tissue fixation (158,159). This procedure appears to reduce the time for tissue sample analysis... 

Ghazani AA, Arneson NC, Warren K, et al. Limited tissue fixation times and whole genomic amplification do not impact array CGH profiles. J. Clin. Pathol. 2006 59 311-315. 

During the tissue fixation process, proteins are cross-linked, causing some epitopes to become undetectable by the staining protocols.10 HIAR reverses this effect, allowing these epitopes to be stained, and therefore has become increasingly important for many IHC staining protocols.19-22 However, the available automated IHC platforms vary in their ability to perform online HIAR. 

If fixative molecules are not present within a tissue, fixation will not occur. 

Jones MD, Banks PM, Caron, BL. Transition metal salts as adjuncts to formahn for tissue fixation (Abstract). Lab. Invest. 1981 44 32A. 

In conclusion, the core components that constitute the mechanism of AR are beginning to come into focus. Further work will be required to solidify these core components and to understand the variations that arise for specific tissues and antigen-antibody pairs. This knowledge, in turn, will lead to improved AR methods, improved ways to overcome the variable effects of tissue fixation, and improved standardization between clinical laboratories. 

Hopwood D, Shdders W, Yeaman GR. Tissue fixation with phenol-formaldehyde for routine histopathology. Histochem. I 1989 21 228-234. 

Our plan for editing this book was enhanced by the Histochemical Society Annual Meeting held at the Experimental Biology 2007 Meeting in Washington, DC. Several interesting workshops with respect to tissue fixation for molecular... 

Hassell, J., and Hand, A. (1974) Tissue fixation with diimidoesters as an alternative to aldehydes. I. Comparison of cross-linking and ultrastructure obtained with dimethylsuberimidate and glutaraldehyde./. Histochem. Cytochem. 22, 223-239. 

The procedures described represent guidelines optimal methods should be determined in individual laboratories. This is because of the variability of tissue fixation and processing and the variety and stability of antigen targets. 

Sometimes it is a good approach to check the quality of tissue fixation at this stage (see Note 24). 

In support of Dr. Shi s opinion is the fact that in some cases the absolute specificity of even monoclonal antibodies can be questioned. The absorption control cannot always determine whether the protein bound in the tissue is the same protein used for absorption. The monoclonal antibody may instead recognize a similar epitope of an unrelated protein, especially following tissue fixation. Absorption controls therefore may not provide the specificity of the antibody for a protein under study in the tissue. 

FIGURE 1.1. Electron micrographs of liver tissue processed differently for transmission electron microscopy. (A) Processed by the conventional method. Note superior quality of ultrastructural preservation compared with that obtained with microwave heating. (B) Rapidly processed by vacuum microwave heating. The whole process from tissue fixation to resin embedding was completed in 2 hr. The quality of ultrastructural preservation is satisfactory. Magnification 6,21 Ox (B). (B) courtesy of Richard T. Giberson. 

Another fixative, Kryofix (E. Merck, Darmstadt, Germany) has been recommended as a replacement for formaldehyde in immunohistochemistry by Boon and Kok (1994). Kryofix is a coagulant fixative containing 50% ethyl alcohol and polyethylene glycol (mol. wt. 300). Both ethyl alcohol and polyethylene glycol diffuse rapidly into the tissue, and the tissue fixation is completed in Kryofix in 90 sec with microwave heating. I do not have personal experience with Kryofix. 

Werner, M., Chott, A., Fabiano, A., and Battifora, H. 2000. Effect of formalin tissue fixation and processing on immunohistochemistry. Am. J. Surg. Pathol. 24 1016-1019. 

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