Freeze-Drying from Tertiary Butanol in the Preparation of Endocardium for Scanning Electron Microscopy

Abstract

The scanning electron microscope appearances and shrinkage of blocks of canine endocardium prepared by freeze-drying directly, by freeze-drying after replacing tissue water with tertiary butanol (2-methyl propan-2-ol) and by critical point drying were compared. All three methods demonstrated endothelial cells which showed nuclear prominences, microvilli and intercellular boundaries. The microvilli varied in size and number from dog to dog but were generally less well defined in specimens freeze-dried from water. Shrinkage due to t-butanol dehydration was significantly less than that which occurred in ethanol in the critical point drying method. Overall the reduction in surface area was significantly less in specimens freeze-dried directly at -65 C (6.8%) than in those dried from t-butanol at -20 C (15.4%) and those prepared bly critical point drying (22.1%). However the amount of shrinkage observed in t-butanol treated tissue was not significantly different from that which was critical point dried. It was not possible to distinguish between comparable samples prepared by these two methods on the basis of their scanning electron microscopic appearances. Thus the relative simplicity and convenience of the t-butanol method, together with its saving of time, its use of standard freeze-drying equipment and the avoidance of ice-crystal artefact justify its consideration as an alternative method of preparing wet biological tissue for scanning electron microscopy.

The Maceration Technique in Scanning Electron Microscopy of Collagen Fiber Frameworks: Its Application in the Study of Human Livers.

“…7) They were then freeze-cracked in liquid nitrogen with a razor blade. 8) They were critical-point-dried using liquid CO2 or freeze-dried in t-butyl alcohol (WHEELER et al, 1975;INOUE and OSATAKE, 1988). 9) The preparations were mounted on specimen-holders for SEM with silver paste.…”

Surface properties and porosity of highly porous, nanostructured cellulose II particles

“…The ice crystals act as templates to self-assemble cellulose particles into the freezing direction . In contrast to that, freeze-drying from tBuOH prevents the formation of ice crystals artifacts (Wheeler et al 1975) and is thus superior with regard to preserving the gel structure (Cai et al 2008). This freeze-drying method was also used to obtain aerogels with surface areas of 284-349 m 2 /g starting from cellulose nanofibrils (CNF) (Sehaqui et al 2011;Saito et al 2011;Nemoto et al 2015).…”

On the presence and function of closed lymphatic stomata in the diaphragm of the golden hamster

“…The possibility of some artifact must also be considered. Biological specimens become reduced in their surface width by as much as 2207o during the histological procedures of SEM, and critical-point drying is largely responsible for such shrinkage (Wheeler et al, 1975). Closed stomata are composed of parquets of lymphatic endothelial cells which are adjoined with various junctional abutments.…”