Paraffin tissue sections embedding

Fixation tissue samples for immuno histochemistry were fixed in 2% paraformaldehyde, 0.25% glutaraldehyde and 3% sucrose buffered with 0.05M phosphate buffer pH 7. After incubation for 2 hours at 25 °C and 63 hours at 5°C the specimens were washed 3 x 20 min. in phosphate buffer pH 7. Dehydration was carried out using series of ethanol washings 50, 70, 80, 96% followed by 3 x in 99% (V2 hr in each). After additional treatment with 2x2 hrs in petroleum ether (shellsol D70k, Q7712) and 2 x 2 hrs in paraffin with 7% beeswax, the samples were embedded in paraffin. Cross sections of 12.5 /im were made on a Supercut 2050 Reichart Jung pyramitome. 

Gill et al.21 Archival FFPE spinal cord tissue both paraformaldehyde-fixed frozen rat spinal cord tissue and paraffin-embedded same tissue To establish an optimal protocol for detection of low-abundance protein (NeuN) in human spinal cord FFPE tissue sections, testing three AR solutions of pH 6, alkaline, and acidic buffer, with three heating conditions 95,100, and 105°C Heating FFPE tissue sections in an alkaline buffer yields most effective AR-IHC staining results. 

Shi SR, Key ME, Kalra KL. Antigen retrieval in formalin-fixed, paraffin-embedded tissues an enhancement method for immunohistochemical staining based on microwave oven heating of tissue sections. J. Histochem. Cytochem. 1991 39 741-748. 

Bataille F, Troppmann S, Klebl F, et al. Multiparameter immunofluorescence on paraffin-embedded tissue sections. Appl. Immunohistochem. Mol. Morphol. 2006 14 225-228. 

Shi S-R, Liu C, Balgley BM, et al. Protein extraction from formalin-fixed, paraffin-embedded tissue sections quality evaluation by mass spectrometry. I. Histochem. 

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... 

Bull JH, Harnden P. Efficient nuclear FISH on paraffin-embedded tissue sections using microwave pretreatment. BioTechniques 1999 26 416-422. 

Shi S-R, Cote RJ, Wu L, et al. DNA extraction from archival formalin-fixed, paraffin-embedded tissue sections based on the antigen retrieval principle heating under the influence of pH. /. Histochem. Cytochem. 2002 50 1005-1011. 

Jin L, Majerus J, Oliveira A, et al. Detection of fusion gene transcripts in fresh-frozen and formalin-fixed paraffin-embedded tissue sections of soft-tissue sarcomas after laser capture microdissection and RT-PCR. Diagn. Mol. Pathol. 2003 12 224-230. 

Part of contents pertaining to the protein-coated beads is reproduced with permission from our article entitled Protein-embedding technique a potential approach to standardization of immunohistochemistry for formalin-fixed, paraffin-embedded tissue sections published in J. Histochem. Cytochem. 2005,53(9) 1167-1170. 

Conti CJ, Larcher F, Chesner J, et al. Polyacrylamide gel electrophoresis and immunoblotting of proteins extracted from paraffin-embedded tissue sections. J. Histochem. Cytochem. 1988 36 547-550. 

In 1991 Shi et al. published their seminal observation that high-temperature incubation of formalin-fixed, paraffin-embedded (FFPE) tissue sections in buffers for short periods led to improved immunohistochemical staining.1 However, more than 15 years later, heat-induced antigen retrieval (AR) remains largely an empirical procedure, requiring the optimization of several critical parameters by trial and error.2,3 Further improvements in AR will require an in-depth understanding of the chemistry of formaldehyde fixation and the molecular mechanism(s) underlying the AR method. 

Igarashi H, Sugimura H, Maruyama K, et al. Alteration of immunoreactivity by hydrated autoclaving, microwave treatment, and simple heating of paraffin-embedded tissue sections. APMIS 1994 102 295-307. 

Cattoretti G, Pileri S, Parravicini C, et al. Antigen unmasking on formalin-fixed, paraffin-embedded tissue sections. J. Pathol. 1993 171 83-98. 

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