Intermediate fidelity

For the study of complex systems or of transient plant behavior, more simplified models can also be used, provided that proper experimental validation is obtained. For macro-modeling of the system, an ad hoc steady-state code was developed on the basis of LIBrary for Process Flowsheeting (LIBPF) technology [31], capable either of integrating the detailed codes for stack simulation of the MCFC-D3S and SIMFC, or of using an intermediate-fidelity model for the stack to improve the calculation performance when required. 

Intermediate fidelity— This type of fidelity is estimated when the results of independent trials are obtained with the same method on identical portions of a sample in the same laboratory with different operators, using different equipment over a specific lapse of time. Intermediate fidelity cannot always be estimated. In the case that interests us, that is, GC-MS coupling, an evaluation of intermediate fidelity requires utilizing at least two GC-MS devices and two operators—not possible in... 

After the response functions are plotted from these analyses, one proceeds with the quality control analysis of the samples. The procedure is again using samples of blank matrices spiked in analytes at concentrations chosen by the user over the whole range of the calibration interval. These samples are analyzed based on the response functions plotted to evaluate the accuracy, repeatability, and intermediate fidelity of the method. The concentrations of the quality controls and the results obtained for cocaine from these samples are given in Table 7.3. 

The dosage of a quality control is considered all the more accurate as the value of A is close to 100%. Table 7.4 gives the determined accuracy for each quality control dosing. Repeatability and intermediate fidelity are estimated for each concentration... 

Having determined the accuracy, repeatability, and intermediate fidelity for the four analytes dosed by the method for each of the five concentrations, the next step is to construct a table that compiles all the results (see Table 7.6). The acceptance criteria of an analytical method are 20% for accuracy, coefficients of variation less than 15% for repeatability, and less than 25% for intermediate fidelity according to the SFSTP. 

Estimation of Repeatability and Intermediate Fidelity for Cocaine at a Concentration of 0.5 ng/mg... 

Accuracy, Repeatability, and Intermediate Fidelity Results from Quality Control Samples... 

The third solution, satisfactory in the present case, consists of conserving the EME analyte in the method by adapting the validation criteria to the results obtained and reducing the interval of dosage for this compound. One can, for example, hx a detection limit at 0.10 ng/mg and tolerances of 20 and 30%, respectively, for repeatability and intermediate fidelity for EME. This is possible if the values remain coherent with the toxicological context (as is the case here) and under the condition that these criteria are specified in the validation procedure. Validation is first of all an internal quality procedure. Consequently, the tolerances relative to validation criteria are determined by the analysts or the quality manager of the laboratory. Table 7.7 compiles results relative to the validation of the analytical method presented as an example. 

Accuracy 20 Repeatability 15 Intermediate fidelity 25 Accuracy 20 Repeatability 20 Intermediate fidelity 30 Accuracy 20 Repeatability 15 Intermediate fidelity 25 Accuracy 20 Repeatability 15 Intermediate fidelity 25... 

The fourth solution is no doubt the best attentively studying the result values of Table 7.6. The accuracy, repeatability, and intermediate fidelity values at the concentration of 0.05 ng/mg are very far from the expected results. This indicates a probable detection problem at that concentration. By erasing the corresponding point in the response function, the slope of the latter will be significantly modified. It is probable that if one uses the new response function for the determination of accuracy, repeatability, and intermediate fidelity values at othCT concentrations, the new values will be compatible with the validation criteria. The interesting point of this solution compared to the third one is that it is possible to retain the acceptance criteria at 15% for repeatability and 25% for intermediate fidelity for EME. 

The kinetic mechanism of aminoacyl-tRNA selection demonstrates that the forward steps of EF-Tu GTPase activation and accommodation are crucial for high fidelity. How can this observation be explained on a structural level. Unfortunately, we have high-resolution structures only of the ribosome prior to A-site binding and of the tRNAs bound to the ribosome after accommodation. Of the intermediate states, only the low-resolution structure of a ternary complex EF-Tu—GTP—aminoacyl-tRNA stalled after GTP hydrolysis has been determined by cryo-EM (Figure These studies revealed that EF-Tu interacts with the... 

In addition to proofreading after formation of the aminoacyl-AMP intermediate, most aminoacyl-tRNA synthetases can also hydrolyze the ester linkage between amino acids and tRNAs in the aminoacyl-tRNAs. This hydrolysis is greatly accelerated for incorrectly charged tRNAs, providing yet a third filter to enhance the fidelity of the overall process. The few aminoacyl-tRNA synthetases that activate amino acids with no close structural relatives (Cys-tRNA synthetase, for example) demonstrate little or no proofreading activity in these cases, the active site for aminoacylation can sufficiently discriminate between the proper substrate and any incorrect amino acid. 

Valuable insights into how DNA polymerases process their substrates were obtained as a result of detailed kinetic studies of the enzymes. Benkovic and coworkers employed rapid quenching techniques to study the kinetics of transient intermediates in the reaction pathway of DNA polymerases [5]. Intensive studies revealed that E. coli DNA polymerase I follows an ordered sequential reaction pathway when promoting DNA synthesis. Important aspects of these results for DNA polymerase fidelity are conformational changes before and after the chemical step and the occurrence of different rate-limiting steps for insertion of canonical and non-canonical nucleotides. E. coli DNA polymerase I discriminates between canonical and non-canonical nucleotide insertion by formation of the chemical bond. Bond formation proceeds at a rate more than several thousand times slower when an incorrect dNTP is processed compared with canonical nucleotide insertion. 

Aminoacyl-tRNA synthetases (aaRSs) compose a family of essential enzymes that attach amino acids covalently to tRNA molecules during protein synthesis. Some aaRSs possess a hydrolytic amino acid editing function to ensure the fidelity of protein synthesis. In addition, aminoacylation can occur by indirect pathways that rely on mischarged tRNA intermediates and enzymes other than aaRSs. Throughout evolution, structural and functional divergence of aaRSs has yielded diverse secondary roles. 

The human immunodeficiency virus type 1 (HIV-1) belongs to the family of positive-stranded, enveloped RNA viruses with a DNA intermediate step (retroviruses). Because of the lack of fidelity of the reverse transcriptase (RT), the replication is error-prone, and the infection is characterized by its quasispecies nature. Antiretroviral treatment with such compounds as zidovudine (AZT), zalcitabine (ddC), didanosine (ddl), stavudine (d4T), and lamivudine (3TC) select for quasispecies variants that are resistant to these compounds (1). The detection of these variants is clinically important because they may affect the outcome of the treatment (2). 

Role of RNA intermediates (Herbert and Rich, 1999). The mutation rate of a genome is likely to increase when genetic information is passed through RNA whether RNA is a viral genome or a retrotransposon because RNA polymerase reaction is neither edited nor subject to post-replicative repair. In addition, hotspots of a genetic chain in RNA retrotransposons can result from nomandom patterns of a decreased fidelity strand transfer to other templates and untemplated extensions. 

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