Correlative microscopy of freeze-dried cells and studies on intracellular calcium stores with imaging secondary ion mass spectrometry (SIMS)

Abstract

Secondary ion mass spectrometry (SIMS)-based imaging techniques have become effective tools for studies of elements and molecules in biological samples. In the current work, a correlative microscopy approach was applied to cryogenically prepared fractured freeze-dried cells for organelle-level imaging of chemical composition using SIMS. A CAMECA IMS-3f SIMS ion microscope was used for studying the effect of microtubule-perturbing agents, specifically nocodazole and taxol, on intracellular calcium stores. The perturbation of microtubules in renal epithelial LLC-PK1 cells resulted in significant loss of total calcium in both the nucleus and cytoplasm. In another study, the stable isotope ⁴⁴Ca was used for imaging the influx of calcium in resting and stimulated LLC-PK1 cells. SIMS imaging of two calcium isotopes, ⁴⁴Ca and ⁴⁰Ca, in the same cell revealed the distribution of calcium influx in the ⁴⁴Ca image and endogenous calcium in the ⁴⁰Ca image. An arginine-vasopressin treatment of cells showed that the Golgi apparatus is sensitive to hormonal stimulation.