Fixation perfusion

Kestens, P.J. (1964). La Perfusion du Foie Isole. Editions Arscia S.A. Bruxelles. Meyer, AJ. Lamers, W.H., Chamulead, R.A.F.M. (1990). Nitrogen metabolism and ornithine cycle fixation. Physiol. Rev. 70, 701-748. 

In vivo receptor labeling offers binding conditions similar to those prevailing in the physiological state and it provides anatomical resolutions compatible with perfusion fixation of target tissue. This technique is... 

The use of distilled formaldehyde, not formalin, which contains alcohol, is recommended. Freshly prepared paraformaldehyde can also be used, especially if large volumes of fixative are needed for perfusion fixation. To prepare an 8% solution of paraformaldehyde, in a fume hood add 2 g of paraformaldehyde (trioxymethylene) powder to 25 mL of deionized glass-distilled water. With constant stirring, heat solution to 60-70°C. Once the solution has reached the proper temperature, continue to stir for 15 min. The solution will be milky. Add one to two drops of 1 VNaOH, with stirring, until the solution clears. A slight miUdness may persist. Cool and filter through Whatman No. 1 filter paper. This solution should be used the same day that it is prepared. 

In the subchronic neurotoxicity study, end points measured are similar to those measured in the acute neurotoxicity study. However, the duration of dosing is 90 days and exposure to the test material is usually via the diet. As for the acute neurotoxicity study, these studies consist of three test groups and a control group. The functional observation battery and motor activity tests are conducted at selected intervals such as weeks 5, 9, and 13, as well as pretest. At test termination, at least 6 animals per group are perfused via the heart with fixative to ensure optimal fixation of nervous tissues for histopathology examination. Nervous tissues examined include brain, spinal cord (various segments), and selected nerves such as the optic, sciatic, tibial, and sural nerves. 

Muscles should be prepared for electron microscopy using standard techniques, including rapid fixation (preferably by perfusion) with glutaraldehyde-based fixatives. 

Y3. Yu, Y., Leng, C. G., Kato, Y., and Ohno, S., Ultrastructural study of glomerular capillary loops at different perfusion pressures as revealed by quick-freezing, freeze-substitution and conventional fixation methods. Nephron 76, 452-459 (1997). 

The method of Pickel et al. (1976) is simple for large tissue fragments if the antigen withstands fixation. Organs or animals are perfused with a warm mixture of 1% GA and 1% paraformaldehyde in 100 mM sodium cacodylate buffer, pH 7.15. Microtome sections are pretreated with Triton X-100, normal serum and hypertonic buffer prior to immunostaining. 

Fig. 4. Electron micrographs of L-Asp-LI and L-GIu-LI in hippocampus CAl from a hypoglycemic rat subjected to perfusion fixation. The tissue was treated with uranyl acetate before embedding in epoxy resin. The figure shows accumulation of immunoreactivities over synaptic vesicle clusters (sv) versus over cytoplasmic matrix (cm) in terminals making asymmetrical synapses on spines (s). Broken lines mark the houndary between the vesicle-rich and vesicle-poor parts of the terminals. Scale bar = 0.2 pm. (Modified from Gundersen et al., 1998.)...

Mice are perfused with 20 ml PBS and 20 ml fixative (Table 11.3C) and tissues immersed in fixative at 4°C for 30 min. Fixation with formaldehyde is quenched in cold 0.15 M triethanolamine (pH 7.5) for 30 min and two 5 min washes in cold PBS. Tissues are dehydrated in graded ethanol and embedded in paraffin. After sectioning and adhering to a coated slide, paraffin is removed by dipping three times 5 min in xylene and twice 5 min in ethanol. 

Kimoto Y, Tohyama M, Satoh K, Sakumoto T, Takahashi Y, Shimizu (1981) Fine structure of rat cerebellar noradrenaline terminals as visualized by potassium permanganate in situ perfusion fixation method. Neuroscience, 6, 47-58. 

Fig. 8.5. Human RBC after radiolabeling, examined by scanning electron microscopy. No sign of morphological alteration is visible. Perfusion fixation, critical point drying, xl500...

Fig. 8.9. Human platelets after the radiolabeling procedure, examined by transmission electron microscopy. Almost no signs of platelet activation, manifested as pseudopodia formation. Perfusion fixation, critical point drying, x 40 500...

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