Precision, instrumental profiles

Precision and Calculation Time. To calculate one point on the instrumental profile we need to calculate a multidimensional integral. For the case where absorption can be neglected this integral reduces to a four-dimensional integral. To estimate how the number of points on the calculation grid affects the precision and calculation time, the calculations of the total instrumental profile were performed for four cases. They are given in Table 6.3. 

Figure 6.20 compares two total instrumental profiles calculated with 50 X 50 X 50 X 50 and 5 x 5 x 5 x 5 points. As is easy to see, it is enough to take only 5 calculation points in each direction to reach a precision of 1 %. The calculation time for this case is about 0.05 s. The calculation time can be decreased still further by taking an unequal number of calculation points in each direction from the line position. 

The MARS comes with a software package, operated via the integrated spill-proof key-pad. The instrument can be connected to an external PC, but this is not required for most common operations. Methods and reaction protocols can be designed as tem-perature/time profiles or with precise control of constant power during the reaction. 

HPLC methods can usually be transferred without many modifications, since most commercially available HPLC instruments behave similarly. This is certainly true when the columns applied have a similar selectivity. One adaptation, sometimes needed, concerns the gradient profiles, because of different instrumental or pump dead-volumes. However, larger differences exist between CE instruments, e.g., in hydrodynamic injection procedures, in minimum capillary lengths, in capillary distances to the detector, in cooling mechanisms, and in the injected sample volumes. This makes CE method transfers more difficult. Since robustness tests are performed to avoid transfer problems, these tests seem even more important for CE method validation, than for HPLC method validation. However, in the literature, a robustness test only rarely is included in the validation process of a CE method, and usually only linearity, precision, accuracy, specificity, range, and/or limits of detection and quantification are evaluated. Robustness tests are described in references 20 and 59-92. Given the instrumental transfer problems for CE methods, a robustness test guaranteeing to some extent a successful transfer should include besides the instrument on which the method was developed at least one alternative instrument. 

The more the precision of the instrument, and the more the points for the time unit in the acquired profile, the better the result of the fitting of experimental data. For this reason instruments with a low measure error and connectable to a computer for the automatic and continous aquisition of data are very much prefered. The UV-Vis spectrophotometer is by far the most used instrument in chemical kinetics. It has a good sensitivity and a good control of the temperature. It is connected or easily connectable to a computer and is available nearly everywhere. The absorbance has a very low dependence on the temperature so that, in the used temperature range, its variation can be neglected during the VTK experiments. 

Intermediate precision is to determine method precision in different experiments using different analysts and/or instrument setup. Similar to that of repeatability, one should evaluate the results of individual related substances, total related substances, and the consistency of related substance profiles in all experiments. The percent RSD and confidence level of these results are reported to illustrate the intermediate precision. 

Optical gene chips dense arrays of oligonucleotides have been successfully applied to detect transcriptional profiling and SNP discovery, where massively parallel analysis is required. However, the fluorescence-based readout of these chips involves not only highly precise and expensive instrumentation but also sophisticated numerical algorithms to interpret the data, and therefore these methods have been commonly limited to use in research laboratories. In this way, thin-film arrays of 14, 20, 25, 48 and 64 electrodes have already been fabricated [12,15,39,40,44,48], using lithographic techniques. Readout systems for these arrays based on electrical detection have also been developed. 

Precision studies can be performed under different conditions, and are strongly influenced by variables such as temperature, source and quality of reagents, reproducibility of reagent delivery, and instrumental noise. Therefore, if all precision studies are done in the same laboratory (intralaboratory study) higher precision is expected in comparison with interlaboratory studies, where several laboratories produce the data used to prepare the method precision profile. 

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