Antigen retrieval sections

Sample Frozen FFPE Section with Antigen Retrieval ... 

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. 

Shi SR, Cote RJ, Yang C, et al. Development of an optimal protocol for antigen retrieval a test battery approach exemplified with reference to the staining of retinoblastoma protein (pRB) in formalin-fixed paraffin sections. J. Pathol. 1996 179 347-352. 

Shi S-R, Cote RJ, Chaiwun B, et al. Standardization of immunohistochemistry based on antigen retrieval technique for routine formalin-fixed tissue sections. Appl. Immunohistochem. 1998 6 89-96. 

Yano S, Kashima K, Daa T, et al. An antigen retrieval method using an alkaline solution allows immunoelectron microscopic identification of secretory granules in conventional epoxy-embedded tissue sections. /. Histochem. Cytochem. 2003 51 199-204. 

Naito I, Ninomiya Y, Nomura S. Immunohistochemical diagnosis of Alport s syndrome in paraffin-embedded renal sections antigen retrieval with autoclave heating. Med. Electron Microsc. 2003 36 1-7. 

Jiao Y, Sun Z, Lee T, et al. A simple and sensitive antigen retrieval method for free-floating and slide-mounted tissue sections./. Neurosci. Methods 1999 93 149-162. 

Kan RK, Pleva CM, Hamilton TA, et al. Immunolocalization of MAP-2 in routinely formalin-fixed, paraffin-embedded guinea pig brain sections using micro-wave irradiation a comparison of different combinations of antibody clones and antigen retrieval buffer solutions. Micros. Microanal. 2005 11 175-180. 

Yamashita S, Okada Y. Application of heat-induced antigen retrieval to aldehyde-fixed fresh frozen sections. J. Histochem. Cytochem. 2005 53 1421-1432. 

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. 

Figure 5.2 Comparison of immunohistochemical staining results among variable periods, 6 h to 30 days, of formalin-fixed, paraffin-embedded human breast cancer tissue (A-N), and cell line MCF-7 sections (O-Bl). All four markers, estrogen receptor (ER) (A-G), CK (cytokeratin cocktail, H-N), Her2/neu (O-U), and MIB-1 (V-Bl), showed comparable positive immunostaining results at +++ level after antigen retrieval. Original magnification x 200. Bar = 50 tm. Reproduced with permission from Shi et al., I Histochem. Cytochem. 2007 55 105-109. See color insert.

Figure 5.3 Diagram depicts the further-designed studies to test our hypothesis with respect to standardization of immunohistochemistry based on the antigen retrieval technique exemplified in a multiple direction to draw a conclusion, (a) Periods of formalin fixation, (b) Variable delay of fixation, (c) Storage of FFPE tissue blocks or sections, (d) Variable thickness of FFPE tissue sections, (e) Other variable conditions of processing FFPE tissue blocks. The stereoscopic frame of a cube represents the reliable limitation of quantitative IFIC demonstrated by serial studies as recommended in the text. Reproduced with permission from Shi et al., J. Histochem. Cytochem. 2007 55 105-109.

Figure 7.9 Appropriate antigen retrieval and immunostaining of peptide controls and tissue sections, stained for HER2. The tissue section on the left has an island of 3+ HER2 tumor, toward the top of the tissue section. The tissue section on the right does not express HER2. Identifying information on the label was removed. See color insert.

Figure 7.10 shows the result from a clinical laboratory that did not perform antigen retrieval correctly. The unfixed peptides at 50 and lOpg/mL stained correctly, but the fixed controls did not. The tissue section (left slide, labeled HER2-99 ) also did not stain well, since it requires antigen retrieval. 

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. 

Brown D, Lyndon J, Tilley SA, et al. Antigen retrieval in cryostat tissue sections and cultured cells by treatment with sodium dodecyl sulfate (SDS). Histochem. Cell Biol. 1996 105 261-267. 

Kakimoto K, Takekoshi S, Miyajima K, et al. Hypothesis for the mechanism for heat-induced antigen retrieval occurring on fresh frozen sections without formalin-fixation in immunohistochemistry. J. Mol. Histol. 2008 39 389-399. 

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