ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.303

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractWe have developed a reliable nonisotopic mRNA in situ hybridization (ISH) technique utilizing specific antisense riboprobe and oligodeoxynucleotide probes labeled with biotin or digoxigenin to detect human glial fibrillary acidic protein and human S-100βprotein in formalin fixed, paraffin embedded human and rat brain tissue sections. This method also allows detection of T and B cell rearrangements with commercially prepared oligodeoxynucleotide probes for kappa and lambda light chains in lymphatic tissues. The method uses proteinase K digestion and overnight hybridization with the labeled probes. Detection of the hybridized probe is accomplished with either streptavidin alkaline phosphatase or with anti-digoxigenin-AP. Hybridization is visualized after enzyme reaction with nitro blue tetrazolium substrate at the light microscopy level. ISH with biotin or digoxigenin labeled oligodeoxynucleotide probes and riboprobes is a rapid, accurate and specific procedure for the study of mRNAs in tissue sections. It correlates well with results obtained by immunohistochemistry. This ISH protocol should be extendible for the detection of other mRNAs for which the sequence is known. (The J Histotechnol17:307, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.307

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractWe report a modified method for non-radioactive in situ hybridization suitable for semiquantitative analysis of specific mRNA sequences in fixed cultured cells or formalin fixed, paraffin embedded tissue sections. Our modifications include random primer incorporation of digoxigenin-label into DNA probe; specimen treatment with proteinase K; heating of specimen and hybridization solution to 100°C for 10 min followed by hybridization at 42°C for 18 hr; and improved immunologic detection with an antibody against digoxigenin conjugate. Visual results were obtained within 3 days. Probe sizes as large as 1.8 kb resulted in good specific signals with minimal background. Specific histologic localization can be semiquantitated under a brightfield microscope linked to a computer aided densitometry system. (The J Histotechnol17:313, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.313

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractA recently described antigen retrieval method with microwave heating of tissue sections was combined with trypsin digestion to increase the immunostaining intensity of estrogen and progesterone receptors. We used 2 new monoclonal antibodies, the estrogen receptor (ER)-specific ER/D5 and the progesterone receptor (PR)-specific PRlOA9, to detect hormone receptors in tissue sections of routinely fixed and embedded breast cancer specimens. These 2 antibodies react with their antigens only following microwave pretreatment of the fixed-embedded sections. When we used trypsin digestion before the microwave heating, the intensity of nuclear hormone receptor immunostaining increased. In some cases, we also found cytoplasmic and/or membrane-associated staining when using the PR-specific PRlOA9 antibody. However, when we used trypsin digestion following the microwave heating, the nuclear staining intensity of ER decreased and that of PR increased. We also observed a loss or significant decrease of the cytoplasmic and membrane-associated components of PR immunostaining. We suggest that using different tissue pretreatment protocols, we can distinguish between the 2 different forms of the 2 steroid hormone receptors. (The J. Histotechnol17:321, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.321

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractThe effects of fixation procedures on the immunoreactivity of PCNA were investigated in smear preparations of HeLa S3 cells. The cells were fixed with acetone, methanol at 4°C, or both, and immunostaining was performed with a monoclonal antibody against PCNA (19A2). Although we failed to detect PCNA with either fixative alone or with a mixture of the 2 fixatives, 15 min fixation with acetone followed by 15 min fixation with methanol at 4°C yielded excellent nuclear staining. First, the PCNA labeling index (LI) was measured and compared with that of BrdU in smear preparations of HeLa cells fixed with acetone followed by methanol. The PCNA LI values (34.7±1.2% , n±10) we every similar to those of BrdU (36.8±0.8% , n = 10). Double immunostaining of PCNA/BrdU was then performed on HeLa cell smear preparations fixed by the same procedure. We observed cells positive for both antibodies with similar frequency of the labeling index values for 2 parameters in the smears. In contrast, there were no cells positive for either parameter alone. These results strongly suggest that, with 15 min fixation with acetone followed by 15 min fixation with methanol at 4°C, PCNA detection is specific for S phase cells. (The J Histotechnol17:325, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.325

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractThe effects of 10 fixatives and formalin fixation time were assessed on the immunoreactivity of KP1 (CD68 antigens) in colorectal tissues in paraffin embedded sections. A panel of fixatives including B5, periodate-lysine-paraformaldehydedichromate, periodate-lysine-paraformaldehyde, 4% paraformaldehyde (PF), zinc formalin, formol dichromate, 10% neutral buffered formalin (NBF), Bouin fluid, Carnoy fluid, and acetone were used to determine an optimal fixative. Another set of colorectal tissues were fixed in 10% NBF progressively at intervals of 1,2,4, and 20 hr; 2,7, and 14 days; and 4 wk to investigate the formalin-induced detrimental effect on CD68 antigens of macrophages.The best result was found after fixation in B5. Short fixation in NBF for 4 hr with trypsinization also gave satisfactory results. There was a distinct decrease in staining intensity after 20 hr exposure to NBF, and the reactivities were totally abolished after 2 days. Trypsinization did enhance the staining intensity significantly with no discrimination of fixatives. (The J Histotechnol17:329, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.329

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractClinical, radiographic, fluorescence, histological, and electron microscopy techniques can be utilized for a comprehensive examination of bone/implant interfaces. To accomplish this objective, proper fixation, processing techniques, instrument quality control, and laboratory safety practices should be followed at all times. This paper reviews the methodology that has become standard practice in our laboratory for the evaluation of bone/irnplant interfaces. A regimen of fluorescent bone markers was used after implant placement to study rates of bone remodeling. At sacrifice, the tissue was perfused or immersed in an aldehyde or alcohol fixative. Implants were retrieved en bloc, radiographs made, and tissues embedded in plastic. Tissue implant blocks were cut longitudinally into 300–400μm thick wafers with a low speed saw equipped with a diamond blade. The wafers were glued to plexiglass slides and hand ground to approximately 25μm. Wafers were photographed with an epi-fluorescence microscope; radiographs were made in an X-ray cabinet; and the sections were stained for light microscopy evaluation. Various histomorphometric methods can be used to determine some variables of tissue remodeling around the implants from photomicrographs. Specimens prepared in this fashion can also be evaluated with a scanning or transmission electron microscope. These combined techniques provide a comprehensive approach for the evaluation of the bone/implant interface. (The J Histotechnol17:333, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.333

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractFor many years, benzoyl peroxide has been used as an initiator for polymerization of methylmethacrylate. In 1983, di(4-tert-butylcyclohexyl) peroxydicarbonate, trade-named Perkadox 16, was introduced in the literature as an alternative to benzoyl peroxide.When Perkadox 16 was incorporated into the protocols in our respective laboratories, we found that it had some unique advantages over benzoyl peroxide, including reduced preparation time and reproducible polymerization. However, during the course of the last 4 years, we have begun to experience some problems. This paper presents some of the problems that have been associated with Perkadox 16 in 2 different laboratories and how each laboratory has worked out a system to avoid them. (The J Histotechnol17:343, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.343

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractA method to identify concurrent viral respiratory infections in animals withPneumocystis Carinii(PC) has been developed by combining immunohistochemical and conventional special staining methods. In an effort to reduce both the number of slides to be screened microscopically and to demonstrate viral expression and PC cysts in the same paraffin section, a streptavidin immunohistochemistry protocol was combined with a rapid Grocott methenamine silver stain. This method is within the expertise and technical capabilities of most research and clinical laboratories. (The J Histotechnol17:349, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.349

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor

摘要:

AbstractA new method for the demonstration of Alzheimer's disease is presented. The method includes pretreatment in a low concentration of uranyl nitrate followed by sensitization in 0.1% silver nitrate for 3 to 5 minutes. Development is done in a solution consisting of hydroquinone, gum mastic, and silver nitrate. This method allows sectioning at 5μm or lower, enabling excellent visualization of the senile plaques and neurofibrillary tangles associated with aging and Alzheimer's disease. The method takes about 20 minutes to perform. (The J Histotechnol17:353, 1994)

ISSN:0147-8885    

DOI:10.1179/his.1994.17.4.353

出版商:Taylor&Francis    

年代:1994

数据来源: Taylor