Name: high precision zinc isotopic measurements applied to mouse organs
Uploaded: 2015-05-22T13:00:00
Description: Watch this Scientific Journal Video about High Precision Zinc Isotopic Measurements Applied to Mouse Organs at JoVE.com

FM acknowledges funding from the ANR through a chaire d&#8217;Excellence IDEX Sorbonne Paris Cit&#233;, the INSU through a PNP grant, the Institut Universitaire de France as well as the Labex UniverEarth program at Sorbonne Paris Cit&#233; (ANR-10-LABX-0023 and ANR-11-IDEX-0005-02). We also thank funding from the European Research Council under the European Community&#39;s H2020 framework program/ERC grant agreement #637503 (Pristine).

NOTE: Procedures involving animals have been approved by the Institutional Animal Care and Use Committee (IACUC) at the Universit&#233; Paris Diderot.
1. Preparation of Materials
<ol>
	<li>Sub-boil distill 1 L of the acids (HNO3, HBr) in order to purify them from impurity.</li>
	<li>Clean the beakers and tip adaptor in a hot (~ 100 &#176;C) concentrated HNO3 acid bath for at least two days.</li>
	<li>Wash the pipette tips in a cold 3 N HNO3 bath for several days and rinse individ...

<ol>
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The reproducibility of the measurements is evaluated through replicated analyses of the same samples carried out during different analytical sessions. For example6, we have replicated the same terrestrial rock 7 times and we obtained the results reported in the Table 2.
As expected from the theory of isotopic fractionation10 and as measured in any solar system material so far (e.g.,&#160;meteorite11-13, plants3-5, deep-sea s...

The measurement of high-precision (better than 100 ppm/atomic mass unit) zinc stable isotope composition has only been possible for about 15 years thanks to the development of multi-collector plasma-source mass-spectrometers and has since been mostly applied in Earth and planetary sciences. The applications to the medical field are novel and have a strong potential as biomarkers for diseases that modify the metabolism of zinc (e.g., Alzheimer disease). This paper reports a method to measure with high precision the natural stable isotope ratios of zinc in various mouse organs. The same would be applicable to human samples. The method consists of the dissolution of the organs, the chemical purification of zinc from the rest of the atoms, and then the analysis of the isotope ratio on a mass-spectrometer.
The quality of Zn isotopic measurements is dependent on the quality of the chemical purification (purity of Zn, low blank compared to the amount of Zn present in the sample, high chemical yield of the procedure) and on the control of the instrumental bias. The high purity of the final Zn fraction is needed to remove both isobaric interferences and non-isobaric interference that create a matrix effect. Isobaric nuclides create direct interferences (e.g., 64Ni). Non-isobaric interferences generate the so-called &#8220;matrix&#8221; effect and alter the analytical precision of the measurements by changing the condition of the ionization compared to the pure zinc standard to which the samples are compared to1. A low blank (&#60; 10 ng) indicates that there is no contamination of the samples by external Zn that would bias the measured isotopic composition. As Zn isotopes can be fractionated during ion-exchange chromatography2, the collection of all the Zn atoms ensures that no isotopic fractionation occurs, which implies that the chemical procedure should have a full yield. Finally, the correction of the instrumental isotopic fractionation during the mass-spectrometry measurement is done via the &#8220;standard bracketing&#8221; method.
Therefore, the main difficulties to obtain precise measurements are controlling the external contamination (i.e., low blank), producing a full yield chemical purification that is clean of any other atoms or molecules, and correcting the instrumental isotopic fractionation on the mass-spectrometer. In this paper we will describe our analytical protocol to separate Zn from the mouse organs as well as the mass-spectrometry measurements.
The extraction is done using a low quantity of diluted acids (HBr/HNO3 media) on micro-columns (0.5 &#181;l and 0.1 &#181;l) of anion-exchange resin. It has a full yield and the measurements have an external reproducibility better than 50 ppm on the 66Zn/64Zn ratio. Another advantage of the method is that it is very fast. The method is therefore very well adapted to medical sciences, in which one needs to analyze a large number of samples compared to geosciences, where these analytical methods were developed.

<table border="0" cellpadding="0" cellspacing="0" width="601">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:236px;">Name of Material/ Equipment</td>
			<td style="width:99px;">Company</td>
			<td style="width:99px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:236px;">Multi-collection inductively-coupled-plasma mass-spectromter</td>
			<td style="width:99px;">Thermo-Fisher</td>
			<td style="width:99px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Anion-exchange resin AG1 X8 200-400</td>
			<td style="width:99px;">Bio-Rad</td>
			<td colspan="2">140-1443-MSDS</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:236px;">teflon beakers</td>
			<td style="width:99px;">Savillex&#160;</td>
			<td>200-015-12</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:236px;">Home-made teflon colunms made with shrinkable teflon</td>
			<td style="width:99px;"></td>
			<td style="width:99px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

high precision zinc isotopic measurements applied to mouse organs

We present a procedure to measure with high precision zinc isotope ratios in mouse organs. Zinc is composed of 5 stable isotopes (64Zn, 66Zn, 67Zn, 68Zn and 70Zn) which are naturally fractionated between mouse organs. We first show how to dissolve the different organs in order to free the Zn atoms; this step is realized by a mixture of HNO3 and H2O2. We then purify the zinc atoms from all the other elements, in particular from isobaric interferences (e.g., Ni), by anion-exchange chromatography in a dilute HBr/HNO3 medium. These first two steps are performed in a clean laboratory using high purity chemicals. Finally, the isotope ratios are measured by using a multi-collector inductively-coupled-plasma mass-spectrometer, in low resolution. The samples are injected using a spray chamber and the isotopic fractionation induced by the mass-spectrometer is corrected by comparing the ratio of the samples to the ratio of a standard (standard bracketing technique).&#160;This full typical&#160;procedure produces an isotope ratio with a 50 ppm (2 s.d.) reproducibility.

In 1.5 N HBr, the main zinc species (ZnBr3-) forms very strong complexes with the anion-exchange resin, while most other elements do not interact with the resin. Zinc is then recovered by changing the medium to diluted HNO3, changing the speciation of Zn to Zn2+ which is released from the resin6,7.
Isotope ratios are typically expressed as parts per 1,000 deviations relative to a standard:
<img alt="Equation 1" fo:content-width="2.5i...

Watch this Scientific Journal Video about High Precision Zinc Isotopic Measurements Applied to Mouse Organs at JoVE.com

High Precision Zinc Isotopic Measurements Applied to Mouse Organs

We present the technique to measure with high precision zinc isotope ratios in mouse organs.

We present a procedure to measure with high precision zinc isotope ratios in mouse organs. Zinc is composed of 5 stable isotopes (64Zn, 66Zn, 67Zn, 68Zn ...

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