Novel epifluorescence microscopy method to determine life position of foraminifera in sediments

Abstract. Microscopic examination of resin-embedded sedimenl cores has provided information regarding the sedimentary fabric in certain environments (e.g. Watling, 1988) as well as the in situ associations of rose Bengal-stained infaunal foraminifera (e.g. Frankel, 1970, 1974). Unfortunately, difficulties connected with making and examining these resin-embedded cores have limited their widespread use in studying meioinfauna. In particular, sediment grains may obscure specimens when viewed by transmitted light microscopy. Also, using rose Bengal to identify live foraminifera can be problematic (Bernhard, 1988). Furthermore, fine-grained deposits are difficult to infiltrate with highly viscous embedding media (e.g. Epon 812 ≥ 100 centipoise). and may require a vacuum apparatus for proper infiltration (Watling, 1988).

Fluorescent probes were recently used to distinguish live from dead foraminifera (Bernhard et al., 1995). Here we report a novel method using an aldehyde-fixable fluorescent probe (Cell Tracker^™ Green CMFDA) which, when viewed by epifluorescence microscopy, greatly enhances the visibility of biological material alive at the time of fixation. Cell Tracker^™ Green CMFDA is a membrane-permeant chloromethyl derivative of fluorescein diacetate that is hydrolyzed by cellular esterases to yield a membrane-impermeant, fluorescent intermediate which further undergoes a reaction with glutathione to form an aldehyde-fixable end-product (Haugland, 1992). We also used low viscosity embedding media (i.e. 20–60 centipoise) to facilitate infiltration. The method is applied to the study of live foraminifera in a sediment core.

PROCEDURE

A c. 1.5 cm diameter subcorer (fashioned from a 10 ml syringe by removing the tip end, creating a cylinder into which two syringe plungers were inserted) was used to . . .