Mounting pollen on a thermoplastic adhesive for scanning electron microscopy

Abstract

A heat-sensitive adhesive, Tempfix?, developed for scanning electron microscopy of powders and small particles, provides consistently high-quality secondary imaging with a variety of pollen grain morphologies and preparation protocols. Tempfix" was used successfully under all combinations of these conditions: (1) pollen air-dried from ETOH or artificially dried; (2) pollen transferred to naked or sputter-coated Tempfix?; and (3) pollen transferred to solid (at room temperature) or tacky (40-43?C) Tempfix?. In contrast to other adhesives, Tempfix": (1) does not react with the dehydration fluid ethanol; (2) permits control of depth of pollen penetration into the adhesive; (3) provides a high pollen retention rate with both air-dried and artificially dried preparations; and (4) has a smooth glassy background. A critical element in many scanning electron microscopic (SEM) studies of pollen is the adhesive used to anchor grains to the specimen mount. Surprisingly, palynological adhesives have received relatively little attention. Frequently, they are unreported or only briefly noted in the materials-and-methods sections of pollen investigations. Adhesives for pollen grains are similar to those for other particulates; they include double-stick tape, rubber cement, silver paste, poly-L-lysine, glues, and waxes (Murphy, 1982). All of these, to some extent, yield inconsistent or undesirable results: charging (the buildup of electrons caused by inadequate sample grounding); sunken grains with halos attributable to partial embedment in the adhesive; grains with obscured surfaces caused by interaction of dehydrating solution with the adhesive; wrinkled, grainy, and uneven backgrounds owing to the nature and composition of the adhesive; and excessive loss of pollen grains caused by inadequate bonding with the adhesive. Recently, a thermoplastic adhesive, Tempfix?, was recommended as a mounting medium for small particles and powders in SEM (Marchese-Ragona et al., 1992). Because Tempfix? offered potential advantages over the adhesives listed above, particularly the ability to control the depth of sample penetration, we tested this adhesive using a variety of pollen grain sculpture patterns and SEM preparation techniques. MATERIALS AND METHODS Dry pollen was collected from herbarium specimens (Table I) and treated as outlined in Fig. 1. First, pollen was acetolyzed (10 min of boiling in a 9:1 mixture of acetic anhydride and sulfuric acid; Erdtman, 1960). All samples then were dehydrated through graded ethanol (ETOH) solutions to 100% ETOH, and then either (1) air-dried directly onto specimen mounts (Fig. 1: method TRANS. AM. MICROSC. Soc., 113(1): 72-79. 1994. ? Copyright, 1994, by the American Microscopical Society, Inc. This content downloaded from 157.55.39.243 on Wed, 05 Oct 2016 04:52:18 UTC All use subject to http://about.jstor.org/terms VOL. 113, NO. 1, JANUARY 1994