Multielement analysis of biological samples by inductively coupled plasma-mass spectrometry. II. Rapid survey method for profiling trace elements in body fluids.

A rapid survey of the elements in biological materials, covering most of the elements in the periodic table, is possible by using available software for semi-quantitative analysis (SEMI-QUANT) by inductively coupled plasma-mass spectrometry. The procedure takes 5 min after sample preparation and gives results with a precision (CV) of approximately 20%. At a 10-fold dilution, 13 elements can be consistently and reliably detected in serum and 15 elements in whole-blood samples. At present the most important limitation of this method is mass overlap by polyatomic species for some elements of interest (e.g., Cr, Mn, and V). However, for the set of elements that can be reliably determined at endogenous concentrations, including Li, B, Mg, Fe, Cu, Zn, Rb, and Sr, the rapid scanning capability may be useful. Although matrix effects limit the direct interpretation of the semi-quantitative output, reasonable estimates of concentration are attainable by using matrix-matched standards or by adding a multielement standard to an aliquot from one sample in the set. We also present an example of determination of 25 elements in saliva from a patient with extensive dental work: Components of many of his dental alloys were readily identified. The method may also prove useful for screening multiple toxic exposures to heavier elements, such as Pb, Tl, Cd, and Hg.

Localization of calcium in roots and microsomal membranes of corn by direct pyroantimonate precipitation.

Many cell membrane systems, including microsomal vesicles of corn, are able to regulate calcium levels both in vivo and in vitro, often in an ATP-dependent, calmodulin-stimulated fashion. The purpose of this study was to determine calcium distribution in meristematic cells of intact tissue and microsomal vesicles from corn roots using direct pyroantimonate-osmium fixation. In root cells, precipitates were localized in mitochondria, plastids, the nucleus, endoplasmic reticulum, Golgi apparatus, and along the plasma membrane. Plasma membrane-enriched microsomal vesicles isolated from corn roots incubated in media to permit calcium transport before pyroantimonate-osmium fixation show internal precipitates associated with the membrane and in the lumen of the vesicles. De-staining of the sections with 1 mM EDTA or EGTA removed precipitate from the sections, confirming the presence of calcium in the antimonate precipitates. These data support biochemical data that this same membrane preparation exhibited ATP-dependent calcium sequestration that was stimulated by calmodulin, as measured by retention of 45Ca. This provides evidence that these membranes are responsible for ATP-requiring, calmodulin-stimulated calcium transport in the intact cell.

Effects of microfilament disrupters on microfilament distribution and morphology in maize root cells.

Maize root tip cells were examined for the distribution of actin microfilaments in various cell types and to determine the effects of microfilament disrupters. Fluorescence microscopy on fixed, stabilized, squashed cells using the F-actin specific probe, rhodamine-labelled phalloidin, allowed for a three-dimensional visualization of actin microfilaments. Microfilaments were observed as long, meandering structures in root cap cells and meristematic cells, while those in immature vascular parenchyma were abundant in the thin band of cytoplasm and were long and less curved. By modifying standard electron microscopic fixation procedures, microfilaments in plant cells could be easily detected in all cell types. Treatment with cytochalasin B, cytochalasin D and lead acetate, compounds that interfere with microfilament related processes, re-organized the microfilaments into abnormal crossed and highly condensed masses. All the treatments affected not only the microfilaments but also the accumulation of secretory vesicles. The vivid demonstration of the effects of all of these microfilament disrupters on the number and size of Golgi vesicles indicates that these vesicles may depend on microfilaments for intracellular movement.

An autoimmune antibody from scleroderma patients recognizes a component of the plant cell nucleolus.

An auto-antibody from human serum of patients with the autoimmune disease scleroderma was used to localize the nucleolus in meristematic cells of onion and soybean roots using indirect immunofluorescence microscopy. Similar lots of antiserum recognized a single 34 kD, nucleolar protein, fibrillarin, in a variety of animal cells (Ochs et al. 1984, 1985). In both plants, antibody linked fluorescence is associated with the one to several nucleoli present in the interphase nucleus. The fluorescence becomes diffuse around condensing prophase chromosomes and becomes more diffused at metaphase with slightly more intense fluorescence surrounding the chromosomes. At anaphase-telophase the fluorescence is localized in dense areas within the chromosomes, presumably representing prenucleolar bodies which will form the interphase nucleoli of the daughter nuclei. This antiserum provides a new, valuable tool for the study of the nucleolus and the highly conversed nucleolar antigen(s) that it recognizes.