Scanning Microscopy


Much of the current knowledge of Pb sequestration in earthworm chloragosome granules stems from quantitative and semi-quantitative Electron Probe X-ray Microanalysis (EPXMA) of unfixed, air-dried tissue smears. Fully quantitative EPXMA was used in this study to assess the validity of smearing by comparison with unfixed, freeze-dried cryosections. Samples were obtained from two earthworm species, Lumbricus rubellus (L.r.) and Dendrobaena rubida (D.r.) collected from a Pb and Zn polluted soil (confirmed by XRF). A number of conclusions emerged :- (a) the element compositions of whole worms, determined by EPXMA of sprayed microdroplets of wet-digested tissues, were different (D.r. : Pb = 29.9, Zn= 33.9, Ca = 207.3, S = 387.9, P = 258.0; L.r. : Pb = 12.3, Zn = 44.4, Ca = 288.1, S = 311.8, P = 354. 9 mM/kg dr.w.); (b) the major proportions of the body burdens of Pb and Zn are located in the chloragogenous tissue; (c) cryosectioned chloragosomes possess a concentric sub-structure, and the chemical composition of individual granules and granule populations is very heterogeneous; (d) smeared and cryosectioned chloragosome preparations revealed significant species-differences in compositional chemistry (e. g., higher Pb and lower Zn in D.r., c.f. with L.r), which reflected their body burdens; (e) the absolute conc. values presented for L.r. cryosectioned granules are systematically higher than equivalent smear data; the overall patterns of the quantitative data obtained from smeared and cryosectioned chloragosomes were substantially similar, suggesting that the simpler technique can be recommended for certain applications, particularly where monitoring a large granule population is paramount.

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