Paraffin tissue sections

Watters AD, Bartlett MS. Fluorescence in situ hybridization in paraffin tissue sections. Mol. Biotechnol. 2002 21 217-220. 

Prior to immunohistochemical staining, paraffin sections must be properly mounted onto slides, and then deparaffinized and rehydrated. To help adherence to the glass and decrease the chances of sections dissociating from the slides, paraffin tissue sections should be mounted on tissue-adhesive-coated slides. The use of tissue-adhesive-coated slides is especially important for paraffin tissue sections undergoing heat-induced antigen retrieval (see Sect. 6.1.1). 

Endogenous biotin may be a cause for nonspecific background staining (see Sect. 5.4). To eliminate this unwanted staining, apply an avidin/biotin block (for instance Avidin/Biotin blocking kit from VECTASTAIN, Cat. No. SP-2001). Usually, paraffin tissue sections are free from endogenous biotin, and this step may be omitted. 

Histology-grade xylene. Use for removing paraffin from paraffin tissue sections on glass slides. 

Suurmeijer, A. J. H., and Boon, M. E. 1999. Pretreatment in a high-pressure microwave processor for MIB-1 immunostaining of cytological smears and paraffin tissue sections to visualize the various phases of the mitotic cycle./. Histochem. Cytochem. 47 1015-1020. 

Double staining can be done however, determination of surface expression of more than one antigen is very difficult cytoplasmic and surface or nuclear and surface antigen expression relatively easy Wide spectrum of antibodies for frozen tissue sections relatively similar as for flow, but sometimes less useful because of sensitivity for paraffin tissue sections less, but with antigen retrieval the gap is narrowing Architecture identifiable and sometimes highlighted by immunostaining... 

In comparison to most other B cell lymphomas, MCL is relatively unique based on overexpression the cell cycle protein cyclin Dl, which can be identified immunophenotypically in the nuclei of the tumor cells in formalin-fixed paraffin tissue sections (B8, D3, K34, S30, S33, V7, Y4, Z4). The overexpression of the cyclin Dl protein is due to the characteristic genetic translocation found in MCL, t( 11 14), which juxtaposes heBCL-l (PRAD1, CYCLIN Dl) gene on chromosome 11 next to the immunoglobulin heavy-chain gene on chromosome 14 (R12, W7, W8). 

The median survival of patients with MCL is 3-5 years. However, if the patients develop additional abnormalities with respect to other cell cycle proteins, such as p53, pi 6, and pi 8, including structural alterations, the median survival decreases to approximately 18 months. Immunodetection of p53, p21, and pl6 (all of which can be identified in paraffin tissue sections) is associated with more aggressive disease and a poor prognosis (L4, P12). In addition, a high proliferation rate, which can be determined by immunostaining for Ki-67, is also associated with a poor prognosis (A13, B21, S32). 

Approximately 70% of PTCLs exhibit aberrant expression of the pan-T cell antigens. The most frequently lost pan-T cell antigen is CD7, which is absent in approximately 50% of cases, whereas the remaining pan-T cell antigens (CD2, CD3, and CD5) are absent in 10-30% of cases (CIO, D19, P6, P10). Most tumors express CD4, with only small percentage of cases expressing the CD4— CD8+, CD4- CD8-, or CD4+ CD8+ phenotype (D19, P6, P10). In paraffin tissue sections, polyclonal CD3 is a reliable marker of T cells, detecting more than 90% of PTCLs (Cl, CIO, C14, Mil). A variable number of cases express cytotoxic proteins however, the number of positive PTCL cases is less than that of either systemic ALCL or T/NK cell lymphomas (B22, Cll, F8, K14). 

Processing of Paraffinized Tissue Sections FOR Immunoperoxidase Staining... 

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. 

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