Laser ablation-ICP-MS assay development for detecting Cd- and Zn-binding proteins in Cd-exposed Spinacia oleracea L.

Polatajko, Aleksandra; Azzolini, Marisa; Feldmann, Ingo; Stuezel, Thomas; Jakubowski, Norbert

doi:10.1039/b703245e

Cited by 38 publications

(43 citation statements)

References 30 publications

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“…In most cases, the gel was ablated directly, but ablation of blotting membranes [26] and elution of the separated proteins from the gel [4] has also been employed.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of gel electrophoresis conditions for the separation of metal‐tagged proteins with subsequent laser ablation ICP‐MS detection

et al. 2009

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Although laser ablation (LA)-ICP-MS has been reported for the determination of metalloproteins separated by gel electrophoretic techniques (GE), systematic studies that define the conditions essential for successful measurements are still scarce. In this paper we present the results of our studies of basic conditions for the effective application of GE-LA-ICP-MS for the separation of metal-binding proteins, focusing on their stability during GE and post-separation gel treatment. The stability of metal-protein complexes (haemoglobin, myoglobin, superoxide dismutase, carbonic anhydrase, transferrin, albumin, cytochrome c) during GE is dependent on the nature of the metal-protein interaction and the principle of separation. We have observed that non-denaturing GE is a suitable separation technique for most metal-protein complexes (e.g. Zn in carbonic anhydrase and Fe in Tf and myoglobin were quantitatively recovered in a spiked liver cytosol), whereas separation by denaturing GE strongly impaired the stability of the complexes. Equally important is the post-separation treatment of the gel to enable successful detection of the metal. LA-ICP-MS requires drying of the gel without loss of protein-bound metal or cracking of the gel. This was successfully achieved using glycerol followed by heating. We demonstrate that staining of the gel prior to LA-ICP-MS using silver or Coomassie blue is not recommended, since most protein-bound metal is lost during the staining procedure. Furthermore it has been shown that only line scanning with a speed of less than 30 microm/s can reliably distinguish between lines 1 mm apart, while raster spot analysis carries the risk of misinterpretation due to contamination in/on inhomogeneous gels.

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“…In most cases, the gel was ablated directly, but ablation of blotting membranes [26] and elution of the separated proteins from the gel [4] has also been employed.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of gel electrophoresis conditions for the separation of metal‐tagged proteins with subsequent laser ablation ICP‐MS detection

et al. 2009

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“…Since a good spatial resolution (few mm in lateral direction and several hundred nm in depth) can be achieved in LA, it is used for spatial mapping investigations such as line scans of e.g. blotmembranes 6 or elemental depth-profiling of coatings.…”

Section: Introductionmentioning

confidence: 99%

Numerical simulation analysis of flow patterns and particle transport in the HEAD laser ablation cell with respect to inductively coupled plasma spectrometry

Lindner

Autrique

Pisonero

et al. 2010

J. Anal. At. Spectrom.

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The present study analyses a specific laser ablation cell, the High Efficiency Aerosol Dispersion (HEAD) cell (see J. Pisonero et al., J. Anal. At. Spectrom., 2006, 21, 922-931), by means of computational fluid dynamics (CFD) simulations. However, this cell consists of different modular parts, therefore, the results are probably of interest for the further development of other ablation cells. In the HEAD cell, the ablation spot is positioned below an orifice in the ceiling of the sample chamber. The particle transport through this orifice has been analysed for a ceiling height of 0.8 mm. The critical velocity for the onset of particle losses was found to be independent on the ejection angle at the crater spot. The deceleration of the particles can be described as the stopping in an effectively steady gas. Particle losses were negligible in this modular part of the cell at the evaluated laser ablation conditions. The transport efficiency through the Venturi chamber was investigated for different sample gas flow rates. In this case, it was found that small particles were predominantly lost at low flow rates, the large particles at higher flow rates. Making use of the simulation results, it was possible to design a modification of the HEAD cell that results in extremely short calculated washout times. The simulations yielded a signal of less than 10 ms, which was produced by more than 99% of the introduced mass.

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“…Several authors have reported studies on metal losses in proteins during GE [55][56][57][58][59]. In most cases, the use of native PAGE instead of denaturing PAGE was proposed as the best option to minimise the possible losses.…”

Section: Analysis Of Heteroatom-containing Proteins By La-icp-msmentioning

confidence: 99%

Laser ablation ICP-MS for quantitative biomedical applications

et al. 2012

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LA-ICP-MS allows precise, relatively fast, and spatially resolved measurements of elements and isotope ratios at trace and ultratrace concentration levels with minimal sample preparation. Over the past few years this technique has undergone rapid development, and it has been increasingly applied in many different fields, including biological and medical research. The analysis of essential, toxic, and therapeutic metals, metalloids, and nonmetals in biomedical tissues is a key task in the life sciences today, and LA-ICP-MS has proven to be an excellent complement to the organic MS techniques that are much more commonly employed in the biomedical field. In order to provide an appraisal of the fast progress that is occurring in this field, this review critically describes new developments for LA-ICP-MS as well as the most important applications of LA-ICP-MS, with particular emphasis placed on the quantitative imaging of elements in biological tissues, the analysis of heteroatom-tagged proteins after their separation and purification by gel electrophoresis, and the analysis of proteins that do not naturally have ICP-MS-detectable elements in their structures, thus necessitating the use of labelling strategies.

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Laser ablation-ICP-MS assay development for detecting Cd- and Zn-binding proteins in Cd-exposed Spinacia oleracea L.

Cited by 38 publications

References 30 publications

Evaluation of gel electrophoresis conditions for the separation of metal‐tagged proteins with subsequent laser ablation ICP‐MS detection

Evaluation of gel electrophoresis conditions for the separation of metal‐tagged proteins with subsequent laser ablation ICP‐MS detection

Numerical simulation analysis of flow patterns and particle transport in the HEAD laser ablation cell with respect to inductively coupled plasma spectrometry

Laser ablation ICP-MS for quantitative biomedical applications

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