Tissue specimens

Mackey EA, Demiralp R, Fitzpatrick KA, Porter BJ, Wise SA, Becker PR and Greenberg RR (1999) Quality assurance in analysis of cryogenically stored liver tissue specimens from the NIST National Biomonitoring Specimen Bank (NBSB). Sci Total Environ 226 165-176. 

Deep tissue specimens obtained during surgical irrigation and debridement should be sent for Gram stain, culture, and sensitivity.3 Imaging Studies... 

Clinical specimens obtained for the recovery of dematiaceous fungi usually do not require extensive processing. If aspirated specimens contain a substantial amount of purulent material, this can be dissolved with N-acetyl-L-cysteine without sodium hydroxide. Tissue specimens and biopsy material should be homogenized in a tissue homogenizer after highly suspicious areas consisting of necrotic, purulent, or caseous material are selectively examined microscopically and inoculated onto isolation media. 

Hamatani K, Eguchi H, Takahashi K, et al. Improved RT-PCR amplification for molecular analyses with long-term preserved formalin-fixed, paraffin-embedded tissue specimens. J. Histochem. Cytochem. 2006 54 773-780. 

Verbeek DH, Smedts F, Wijnen-Dubbers CW, et al. Histologic processing of thick tissue specimens from cytology slides. Acta Cytol. 1996 40 1198-1204. 

Bratthauer GF. Processing of tissue specimens. Methods Mol. Biol. 1999 115 77-84. 

A reasonable objection to any in vitro model is whether it accurately mirrors the actual process. A strength of this model is that the peptides in the array, mounted on the microscope glass slide, are the very same as the antibody epitopes in the native proteins. Therefore, the types of formaldehyde-induced chemical reactions at or near the epitope are the same as would likely occur in a tissue sample. An additional strength of the model is that the experimental data using the peptide array completely account for the loss of immunoreactivity after formalin fixation and the recovery of immunoreactivity after antigen retrieval (Fig. 16.5). Nonetheless, our data do not prove that the model accurately represents formaldehyde reactions in tissue specimens. For example, our data do not exclude other causes of steric interference. 

Note because of a dramatic increase in the sensitivity this method may require an additional blocking step for inactivation of endogenous biotin in some tissue specimens (see Sect. 5.4). 

Tissue processing tissue specimen (0.5 1.0 mm3) are fixed in 4% buffered formalin for 30 60 min, post-fixed in 1% osmium tetroxide in cacodylate or phosphate buffer, pH 7.2 7.4, stained en bloc for 30 min with 2% aqueous uranyl acetate, then dehydrated in ethanol and embedded in epoxy or acrylic resin. 

Ethical issues as well as difficulty in obtaining enough human nasal tissue specimens have called for the need to use alternative in vitro and in vivo methods. Various in vivo animal models and in vitro excised tissue models have been described in the literature for nasal drug transport studies. However, due to the difficulty in both controlling the experimental conditions in in vivo animal models and obtaining intact excised tissue samples, in vitro cell culture models are also being actively developed. 

Figure 11.1 Ultrastructure of the human lung alveolar barrier. The tissue specimen is obtained via lung resection surgery. (A) Section through a septal wall of an alveolus. The wall is lined by a thin cellular layer formed by alveolar epithelial type I cells (ATI). Connective tissues (ct) separate ATI cells from the capillary endothelium (en) within which an erythrocyte (er) and granulocyte (gc) can be seen. The minimal distance between the alveolar airspace (ai) and erythrocyte is about 800-900 nm. The endothelial nucleus is denoted as n. (B) Details of the lung alveolar epithelial and endothelial barriers. Numerous caveolae (arrows) are seen in the apical and basal plasma membranes of an ATI cell as well as endothelial cell (en) membranes. Caveolae may partake transport of some solutes (e.g., albumin). (C) ATII cells (ATII) are often localised in the comers of alveoli where septal walls branch off. (D) ATII cells are characterised by numerous multilamellar bodies (mlb) which contain components of surfactant. A mitochondrion is denoted as mi.

Two DNA adducts of 1,2-dibromoethane metabolites have been found in in vitro studies (Bolt et al. 1986 Ozawa and Guengerich 1983 Peterson et al. 1988). These adducts are S-[2- (N -guanl)ethyl] glutathione and S-[2-N -guanyl)ethyl] cysteine. These adducts are potential biomarkers of exposure to 1,2-dibromoethane and could be tested for in biopsy or autopsy tissue specimens. 

Emmert-Buck MR, et al. 2000. Molecular profiling of clinical tissue specimens feasibility and applications. Am J Pathol... 

Once the pellet is embedded in paraffin, it can be treated as any tissue specimen would be treated (see Chapter 12). 

The fixative used is again a matter of preference. However, the main reason for preparing this type of specimen is usually to compare fresh cells to the cells found in tissue specimens. Therefore, it is recommended that formalin fixation be used, since this is the fixative of choice for most tissue protocols, and the crosslinking that develops helps to hold the cells together as a pellet. Specimens can be fixed in 95% ethanol/5% glacial acetic acid but will be more fragile in pellet form. 

Experimental samples are mainly derived from tissue culture cells, laboratory animals, or human tissues collected from hospitals after surgical biopsies and autopsies. With human and animal tissue specimens, it is important to arrest metabolic processes within 5-10 min of collection in order to preserve mRNAs from degradation by internal enzymatic reactions (26,27). Most hospitals use 10% buffered formalin as a tissue fixative. Subsequently, each tissue slice is trapped in a paraffin block. Series of 4-5-pm-thick sections are cut and mounted on silanated slides. Formalin-fixed archival tissues have been successfully used in in situ PCR and in situ hybridization protocols (28-32). However, the procedure for RNA protection is not always followed. It is often difficult to alter or control the routine procedures of hospitals for the required protection of mRNAs in surgically removed human tissues. 

Under the Toxics Substances Control Act, the Environmental Protection Agency (EPA) is mandated to gather data on the exposure of the general population to toxic substances. Toward this end, the Office of Toxic Substances within the EPA has undertaken several long term monitoring programs. These programs involve the collection of human tissue specimens from a statistically representative... 

The data needed to meet these objectives are generated on a annual basis by collecting and chemically analyzing adipose tissue specimens for selected toxic substances, mainly organochlorine compounds and polychlorinated biphenyls (PCBs). The 20 compounds... 

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