Cross-linking retrieval

Rait VK, Xu L, O Leary TJ, et al. Modeling formalin fixation and antigen retrieval with bovine pancreatic RBase A II. Interrelationship of cross-linking, immuno-reactivity, and heat treatment. Lab. Invest. 2004 84 300-306. 

These initial findings do not exclude other possible formaldehyde-induced reactions with tissue proteins. Notably, this first model system was not designed to detect the role of lysine residues. Lysine has a propensity to react with and form a variety of different types of cross-links with other amino acids in the presence of formaldehyde.1,3 417 Therefore, it is likely to also be important in reactions with formaldehyde. In fact, peptides with internal lysine residues were purposefully excluded from this initial study for technical reasons. To explore the importance of lysine residues in antigen retrieval, an alternative method was employed. 

Figure 16.5 Immunostained peptide arrays after various treatments of fixation, protein cross-linking, and antigen retrieval, as indicated at the top. Each row has a different peptide that is immunoreactive for the antibody denoted to the left. Column A represents the baseline condition, without any treatment whatsoever. Column B shows immunoreactivity of each peptide after overnight formalin fixation. Column C shows the immunoreactivity after first coating the array with an irrelevant protein (casein) followed by overnight formalin fixation. Column D illustrates the immunoreactivity of the peptides after the treatment of column C, and then antigen retrieval. Reproduced with permission from Reference 15, 2006 American Society for Clinical Pathology.

Two retrieved cross-linked UHMWPE acetabular cups were preliminary investigated by Raman microprobe spectroscopy with respect to their oxidation... 

Relaxation curves can be measured by simple NMR experiments. On the other hand, information about cross-link density and chain stiffness can be retrieved by exploiting... 

NOTE Formalin does not preserve tissue proteins by coagulation but it is thought to form cross links with basic amino acids. Ethanol and mercuric chloride-based fixatives are based on coagulation. With few exceptions retrieval should not be performed on ethanol fixed tissues. It should only be conducted with limited controlled protocols in mercuric-chloride-based fixatives. 

---