Antigen retrieval (AR) incorporating high-temperature microwave (MW) heating of tissue sections before immunostaining is a revolutionary technique that can unmask the antigens in formalin-fixed tissue sections, thus making them available for immunohistochemical staining. Although high temperature is believed to be the primary mechanism in retrieval of antigens, a variety of chemical solutions have been tested to define an optimal AR solution. We tested the hypothesis that pH of the AR solution may influence the quality of immunostaining by using seven different AR buffer solutions at a series of different pH values ranging from 1 to 10. We evaluated the staining of monoclonal antibodies to cytoplasmic antigens (AE1, HMB45, NSE), nuclear antigens (MIB-1, PCNA, ER), and cell surface antigens (MT1, L26, EMA) on routinely formalin-fixed, paraffin-embedded sections under different pH conditions with MW heating for 10 min. The intensity of immunostaining was graded in a blinded fashion. The pH value of the AR buffer solution was carefully measured before, immediately after, and 15 min after the AR procedure. The influence of pH on AR immunohistochemical staining can be summarized into three patterns. Some antigens (L26, PCNA, AE1, EMA, and NSE) showed excellent retrieval throughout the pH range. Other antigens (MIB1 and ER) showed strong intensity of immunohistochemical staining at very low pH and at neutral to high pH, but a dramatic decrease in the intensity of the AR immunostaining at moderately acidic pH (pH 3-6). Still others (MT1 and HMB45) showed increasing intensity of the AR immunostaining with increasing pH, but only weak immunostaining at low pH. Among the seven buffer solutions at any given pH value, the intensity of AR immunostaining was very similar. However, Tris-HCl buffer tended to produce better results at higher pH, compared with other buffers. Although high-temperature heating is believed to be the most important factor for the AR technique, the pH value of the AR solution is an important co-factor for some antigens. Optimization of the AR system should therefore include optimization of the pH of the AR solution. Our results indicate that AR immunostaining of Tris-HCl or sodium acetate buffer at pH 8-9 may be suitable for most antigens, although certain nuclear antigens show optimal staining at low pH.

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BACKGROUND. The expression of E-cadherin in the intercellular adhesion of metastatic prostate cancer cells in bone, which is the most prevalent site of metastatic growth, remains elusive. METHODS. The aim of the study was to compare the concurrent membranous expression of E-cadherin and b-catenin proteins, the state which is known to be associated with the cellular adhesion function of E-cadherin, in prostate biopsy tissue specimens by immunohistochemical staining method. The expression patterns of E-cadherin or b-catenin were classified as homogeneous (most cells exhibiting positively), heterogeneous (a few scattered patches of cells with positivity) or negative. RESULTS. Benign prostate hyperplasia cells exhibited homogeneous expression of both E-cadherin and b-catenin in 9 of 11 (82%), whereas the primary prostate cancer cells were homogeneously positive for both proteins only in 4 of 22 (18%) of the cases. The results are similar to those reported in literature. However, in contrast to the primary cancer, a significantly increased frequency of the metastatic prostate cancer cells in bone exhibited homogeneous expression of E-cadherin and b-catenin in 12 of 17 (71%) of the cases. A statistically significant association was observed between the overexpression of both proteins and the metastatic prostate cancer cells in bone (Fisher's exact P < 0.001). CONCLUSIONS. The result of the study demonstrated for the first time that the membranous overexpression of E-cadherin and b-catenin are significantly associated with the metastatic prostate cancer cells in bone and that the high frequency of expression suggest their involvement in the intercellular adhesion of the metastatic cells in bone.

The study clearly demonstrated a significant association of the concurrent expression of unmethylated E-cadherin gene and E-cadherin protein with metastatic prostate cancer cells in bone, and that its expression may have a role in the intercellular adhesion in the formation of metastatic lesions in bone.

A series of isatin-3-anils (with or without a N-piperidino/morpholinomethyl substituent) have been screened for their cysticidal activity against Schizopyrenus russelli. Their ability to cause excystment has also been studied.