Optimisation of a laser ablation cell for detection of hetero-elements in proteins blotted onto membranes by use of inductively coupled plasma mass spectrometry
Optimisation of a laser ablation cell for detection of hetero-elements in proteins blotted onto membranes by use of inductively coupled plasma mass spectrometry
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DOI：
10.1039/b606773e
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发表时间：
2006-09
影响因子：
3.4
通讯作者：
I. Feldmann;C. Koehler;P. Roos;N. Jakubowski
中科院分区：
文献类型：
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作者：
I. Feldmann;C. Koehler;P. Roos;N. Jakubowski
为了成像常规尺寸膜上蛋白质斑点的元素分布，我们研究了三种不同的激光烧蚀细胞几何形状，以找到一种细胞设计，该细胞在整个细胞中显示出持续的高信号强度，并且具有较短的冲洗时间。为此，对优化后的强度（载气流速、氩气补充气、发电机正向功率）进行了测量，包括在金属钢板上不同位置的单次激光照射，以及在电池线性平移期间连续烧蚀硝化纤维素（NC）膜。在后一个实验中，墨棒结构被打印在NC膜上以模拟斑点蛋白，并用于评估局部分辨率和确定适当的翻译速度。最终细胞的体积约为11.3 cm3，其中单次射击信号在不到一秒钟的时间内被冲洗掉，因此高达1.5 mm s - 1的翻译速度可以应用于距离小于2mm的基线分离结构，这将使用一个由SDS-PAGE分离的磷酸化蛋白混合物的例子进行演示，并将其印迹到NC膜上。
For imaging of elemental distribution in protein spots blotted onto membranes with a conventional size, we have investigated three different laser ablation cell geometries to find a design of a cell showing a constantly high signal intensity across the cell and possessing short wash-out times. For this purpose the resulting intensity after optimisation (He carrier gas flow rate, argon make-up gas, generator forward power) was measured for single laser shots at different positions on a steel metal sheet and for continuous ablation of nitrocellulose (NC) membranes during linear translation of the cell. In the latter experiment, ink bar structures have been printed on the NC membranes to simulate blotted protein spots and used to evaluate the local resolution and to determine an adequate translation velocity. The final cell has a volume of about 11.3 cm3 in which single shot signals are washed out in less than a second so that translation velocities of up to 1.5 mm s−1 can be applied to baseline separate structures with a distance of less than 2 mm, which will be demonstrated using an example of a phosphoprotein mixture separated by SDS-PAGE and blotted onto a NC membrane.