An improved method for perfusion fixation of neural tissues for electron microscopy

Williams, Terence H.; Jew, Jean Y.

doi:10.1016/0040-8166(75)90015-4

Cited by 59 publications

(16 citation statements)

References 8 publications

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“…In the second method, the animal is artificially ventilated during the vascular rinsing and the first few seconds of perfusion with the fixative solution. A simple artificial respiratory method described by Williams and Jew (1975) consists of delivery of lowpressure Carbogen through a plastic tube (8-10 mm inner diameter) that is connected to a thin (1.8-2.5 mm outer diameter), short (4-5 cm) cannula inserted in the trachea of a tracheotomized animal. Closing and opening a hole on the side of the plastic tubing simulates inspiration and respiration.…”

Section: B Vascular Rinsingmentioning

confidence: 99%

Electron Microscopic Preembedding Double-Immunostaining Methods

Leranth

Pickel

1989

Neuroanatomical Tract-Tracing Methods 2

197

242

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Section: B Vascular Rinsingmentioning

confidence: 99%

Electron Microscopic Preembedding Double-Immunostaining Methods

Leranth

Pickel

1989

Neuroanatomical Tract-Tracing Methods 2

197

242

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“…A proper fixation protocol is crucial, as well for the studies on histomorphometric evaluation of peripheral nerve fibers particularly concerning the quantitative features such as axon number, axonal cross-sectional area and 2,4 . Most commonly used fixation protocols in these studies are immersion and perfusion fixation techniques 5,14,16 . Although transcardial perfusion has been approved as the most effective method it indeed has several disadvantages.…”

Section: Discussionmentioning

confidence: 99%

Comparison of Fixation Methods for Peripheral Nerve Fiber

Ulkay¹,

Aktaş²,

Aksoy³

et al. 2013

Kafkas Univ Vet Fak Derg

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Accurate fixation is a must for the assessment of myelinated nerve fiber morphology and transcardial perfusion and immersion methods are the most commonly used fixation techniques. In the present study we designed a new fixation technique for the histomorphometric and stereological evaluation of sciatic nerve fiber and referred it as instillation fixation. The method involved preliminary in situ fixation of the nerve sample without dissecting it from the animal body followed by a complete conventional fixation protocol. The objective of this study was to compare the three fixation techniques with each other for fixation artifacts. Eighteen female Wistar albino rats constituted the study material. The animals were allocated into three experimental groups corresponding to different fixation methods (immersion, instillation and transcardial perfusion, respectively). Quantitative assessments of nerve samples harvested from the animals of each group included the number of total myelinated axons, normal myelinated axons, alterations in myelin compaction, myelinated axons with irregular fiber shape, and with myelin loops and g ratio. Results revealed that normal myelinated axons were markedly lower in the immersion fixation group compared to those of instillation and transcardial perfusion. Moreover a significant decrease was noted with respect to alterations in myelin compaction in the instillation fixation group. In contrast, no significant difference was observed in myelin thickness and axon cross sectional area. In conclusion instillation fixation technique proved to be a valid and simple method for the assessment of peripheral nerve morphology for further analyses in a rat model. Keywords: Rat, Sciatic nerve, Fixation method Periferal Sinir Tespit Yöntemlerinin KarşılaştırılmasıÖzet Doğru bir tespit işlemi miyelinli periferik sinirlerin morfolojik değerlendirmesi için ön koşuldur. Kalp perfüzyonu ve daldırma en yaygın kullanılan tespit yöntemlerindendir. Çalışmamızda bu metotlara ek olarak damlatma tekniği olarak adlandırılan yeni bir tespit metodu uygulandı. Sinir doku örneği canlı hayvan üzerinden çıkarılmadan ön tespit işlemine tabi tutulduktan sonra rutin tespit işlemleri tamamlandı ve üç tespit yönteminin tespit artefaktları açısından birbirleriyle histomorfometrik ve stereolojik olarak karşılaştırılması gerçekleştirildi. On sekiz adet dişi Wistar albino sıçan kullanıldı. Hayvanlar sırasıyla daldırma, damlatma ve kalp perfüzyonu olmak üzere üç eşit gruba ayrıldı. Her bir gruptaki hayvanlara ait siyatik sinir örneklerinin kantitatif değerlendirilmesi toplam miyelinli akson sayısı, normal miyelinli akson sayısı, miyelin kompaksiyonundaki değişimler içeren aksonların sayısı, düzensiz demet yapısı ile kıvrımlı miyelin halkaları içeren miyelinli aksonların sayısı ve g oranları hesaplanarak gerçekleştirildi. Elde edilen bulgulara göre daldırma tespit grubundaki normal miyelinli akson sayısında, damlatma ve kalp perfüzyonu grupları ile karşılaştırıldığında belirgin bir düşüş belirlendi. Bununla beraber damlatma tesp...

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“…Prior work in the central nervous system has shown that immersion can create problematic artifacts in brain tissue, which has unique characteristics that make fixation difficult (Adickes et al, 1997;Beach et al, 1987;Berod et al, 1981;Bondonna et al, 1977;Miller, 1998;Roberts et al, 1990;Simmons et al, 1996;Williams and Jew, 1975). One important reason for the use of perfusion fixation with central nervous system tissue is to minimize anoxia and mechanical damage, since the bony encasements of the skull must be dissected to expose the superficial regions of the brain.…”

Section: Introductionmentioning

confidence: 97%