Paraffin-Embedded Tissues

Faolain et al. deparaffinized parenchymal tissue sections by immersing in two baths of xylene for 5 and 4 min, respectively, followed by two baths of absolute ethanol for 3 and 2 min and a final bath of industrial methylated spirits 95% for 1 min. This method was found through Raman microspectroscopy to be inefficient at removing all of the paraffin since a number of strong signals from C-C and CH2 vibrational modes were observed.  

A recent study, by Meuse and Barker, used FTIR spectroscopy to assess paraffin removal based on CH stretching bands near 2850,2918 and 2956cm. Hexane, xylene and limonene were compared as dewaxing agents and two human breast cancer cell lines were used as model tissues. With all three dewaxing agents, at least 97% of paraffin was found to be removed, with xylene found to be the most effective. The difference from the study by Faolain et is more than likely due to the fact that cell cultures were used, which would not have the complex intracellular components of tissue samples. 

It may be argued that, for FTIR and Raman studies, the removal of paraffin is not necessary at all as only discrete frequency ranges corresponding to the lipid hydrocarbon modes are affected. 

However, visualization of the unstained tissue s anatomical features has been shown to be severely hampered if the paraffin is not removed. Sahu et al. reported that colonic crypts in a 10 pm paraffin embedded tissue section appeared as circular entities when viewed under light microscopy.Moreover, even between adjacent microtomed sections, tissue components can vary 

Drying Decrease in intensity of the 930 cm peak, and an increase in the lipidiprotein signal  

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EXTRACTION OF DNA/RNA FROM FORMALIN-FIXED, PARAFFIN-EMBEDDED TISSUE BASED ON THE ANTIGEN RETRIEVAL PRINCIPLE... 

A simple and effective AR technique of boiling archival paraffin-embedded tissue sections in water to enhance the signal of IHC was developed to circumvent the deleterious effects of formalin fixation, which had previously... 

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Figure 5.1 Diagrammatic explanation of standardization of IHC via AR and test battery to achieve a maximal retrieval level by an optimal protocol of AR. The intensity of IHC (axis y) is inversely correlated with the time of formalin fixation (axis x) as indicated by a reduced slope. Three arrows indicate a potential maximal retrieval level that may equalize the intensity of IHC to a comparable result for routinely processed, paraffin-embedded tissues with various time of fixation. Reproduced with permission from Shi et alHistotechnol. 1999 22 177-192.

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