Fixation techniques

Leonard, J. B. and Shepardson, S. P. (1994) A comparison of heating modes in rapid fixation techniques for electron microscopy. J. Histochem. Cytochem. 42,383-391. 

Improper fixation and processing Proper antigen fixation is the cornerstone of immunoperoxidase techniques, for without it, results will be poor. Some markers may be destroyed by certain fixatives overfixation may mask certain antigens (see Chapter 8) (4). Review manufacturer s package inserts for appropriate fixation techniques. Paraffin-embedded tissue should never be exposed to temperatures >60°C as this can destroy some antigens. 

The replacement of classical textile printing techniques by digital printing techniques (ink-jet and bubble jet) is in full progress. Present limitations result from the availability of appropriate formulations of inks/dyes and fixation techniques. The comparable low production speed and limitations with regard to the quality of the textile material can be expected to be overcome within the next 5-10 years. 

Generally the larger the specimen the longer the time required for fixation, e.g., rabbit heads. There are alternative fixation techniques that may be used, e.g., fixation in Davidson s prior to fixation in Bouin s (4). 

Fox, N. E., and Demaree, R. S. 1999. Quick bacterial microwave fixation technique for scanning electron microscopy. Microsc. Res. Tech. 46 338-339. 

Comparisons have been made of HA localization in skin sections fixed with acid-formalin/ethanol and conventional formalin fixation. Much of the HA, particularly in the epidermis, is eluted during the process of formalin fixation. This suggests that epidermal HA is more loosely associated with cell and tissue structures than is dermal HA. A further incubation of 24 h in aqueous buffer further increases the disparity between the acid-formalin/alcohol and the conventional fixation technique. Once the tissue has been exposed to the acid-formalin/alcohol, the HA association with tissue becomes tenaciously fixed, with little loss of apparent HA observed following additional aqueous incubation, while the formalin-fixed tissues demonstrate progressive loss of HA. 

Weibel, E. R. (1984) Morphometric and stereological methods in respiratory physiology including fixation techniques, in Techniques in Respiratory Physiology, Part 1. (Otis, A. B., ed.), Elsevier, New York, NY, P4/I, P401/1-P401/35. 

The plant was designed to operate the two reactors independently. This allowed a comparison of different catalysts, fixation techniques, hydrodynamics and effluents/model compounds under identical conditions. The plant could be operated with suspended and fixed catalysts. A sedimentation tank was connected to the reactors (via a storage tank) to separate and recycle the catalyst. The experiments could be carried out in continuous or recycling mode [264],... 

Use hypotonic treatment/fixation technique, and affix cells to slide Treat cells with RNase (and/or proteinase K)... 

It is unfortunate that the only approach by which we can currently study cubic membrane structure is from electron microscopy, which often induces artefacts. On the other hand it is more likely that perturbations caused by EM preparation md fixation techniques are destructive to cubic membranes rather than constructive. Therefore it is unlikely that EM preparation techniques would induce the formation of cubic membranes. Indeed similar conclusions have been deduced by several authors dealing with the membrane assemblies that we identify as cubic membranes (see, e.g. [4] and [6]). Besides artefactual problems, our approach is inherently limited, because... 

TABLE 21.1 Antibodies Used with Post-Fixation Technique ... 

Some patients with lymphocytic thyroiditis have antibodies directed against the microsomal or cellular fractions only. Others have positive results for the thyroglobulin antibody, but have no antibodies against the microsomal fraction. The microsomal antibody is the most significant in relation to the etiology of thyroiditis as it is cytotoxic to cultured thyroid cells (D6). Microsomal autoantibodies are detected either by complement fixation technique (CFT) or by immunofluorescence. Be-... 

Comparison of Fixation Techniques. For microwave fixation, fresh tissues were trimmed to approximately 2 mm thickness, placed in plastic cassettes, and held in ice cold saline (30-120 minutes) until irradiation in a 700-watt oven set to hold at 60°C for 2 minutes (7). Microwave-fixed tissues were stored overnight in 70% ethanol at room temperature, processed routinely and then embedded in paraffin, sectioned, deparaffinized, and rehydrated using standard techniques. Other pieces of liver were similarly trimmed, then fixed by immersion in either 3.7% formaldehyde in neutral phosphate buffer (48 hours), 2% glutaraldehyde in phosphate buffer (2 hours), Bouin s solution (24 hours), or B-5 solution (4 hours). These tissues were similarly embedded in paraffin then sectioned routinely. In other studies, specimens of lung and kidney were fixed by microwave irradiation with the same schedule as liver. 

S. Knecht, C. Erggelet, M. Endres, M. Sittinger, C. Kaps, E. Smssi, Mechanical testing of fixation techniques for scaffold-based tissue engineering grafts, J. Biomed. Mater. Res. B Appl. Biomater. 83 (2007) 50-57. 

Sample preparation in electron microscopy is usually performed using chemical or thermal fixation techniques for sample immobilization. The evaluation and details of each preparation technique are presented below. 

Thermal fixation techniques are preferred in the study of microstructured liquids, hydrated biological specimens, fluid-filled porous media, etc. Because thermal fixation is used for such diverse applications, there are a wide variety of fixation techniques and associated equipment that impart microscopic stability to the specimen. 

The essence of cryomicroscopy lies in the ability to vitrify the sample using the thermal fixation technique that alters the sample microstructure to the least extent. Cooling rate estimates of 10 K/s [14], 10 K/s [15], and 10 K/s [16] are quoted as necessary to vitrify water or dilute aqueous suspensions. The ability to achieve the desired cooling rate depends on the sample, the cryogen, and the fixation technique. 10 K/s is probably the most accurate estimate. This evaluation is based on estimates of the cooling rate in freezing techniques that are known to produce vitreous specimens. 

There are basically only two protocols that we use for in situ hybridization to whole-mount tissues, and in essence they are identical one is for whole-mount specimens that can be handled in Eppendorf tubes and the other is adapted for whole-mount specimens on slides. As described above, the challenge in extending these methods to new tissues is to find isolation and fixation techniques that enable them to go through one of these procedures. 

Heat fixation Technique in which air-dried smears are passed through an open flame so that organisms are killed, adhere better to the side, and take up dye more easily. 

---