Intralaboratory

Before a procedure can provide useful analytical information, it is necessary to demonstrate that it is capable of providing acceptable results. Validation is an evaluation of whether the precision and accuracy obtained by following the procedure are appropriate for the problem. In addition, validation ensures that the written procedure has sufficient detail so that different analysts or laboratories following the same procedure obtain comparable results. Ideally, validation uses a standard sample whose composition closely matches the samples for which the procedure was developed. The comparison of replicate analyses can be used to evaluate the procedure s precision and accuracy. Intralaboratory and interlaboratory differences in the procedure also can be evaluated. In the absence of appropriate standards, accuracy can be evaluated by comparing results obtained with a new method to those obtained using a method of known accuracy. Chapter 14 provides a more detailed discussion of validation techniques. 

Standard reference materials provide a necessary but insufficient means for achieving accuracy and measurement compatibiUty on a national or international scale. Good test methods, good laboratory practices, well-qualified personnel, and proper intralaboratory and intedaboratory quaUty assurance procedures ate equally important. A systems approach to measurement compatibiUty is ikustrated in Figure 2. The function of each level is to transfer accuracy to the level below and to help provide traceabiUty to the level above. Thus traversing the hierarchy from bottom to top increases accuracy at the expense of measurement efficiency. 

All the analytical data are from the same laboratory consequently, interlaboratory analytical variation is not a factor. The intralaboratory variation for that laboratory was 9.1 percent (i.e., the relative standard deviation based on repetitive analyses of performance evaluation samples). 

The data presented In Figures 2, 3, and 4 are from five different laboratories However, the samples from any one method In each segment were sent to the same contractor thus. Interlaboratory analytical variation Is not a factor In the data columns, but It may be a factor In the data rows The estimated Interlaboratory and Intralaboratory relative standard deviations for TCDD concentrations of 2 to 12 ppb range from 9 to 18 percent and 5 to 13 percent, respectively ... 

The results of local tissue irritation tests are subject to considerable variability due to relatively small differences in test design or technique. Weil and Scala (1971) arranged and reported on the best known of several intralaboratory studies to... 

In general terms, the variation from laboratory to laboratory (between-laboratory) was greater than that attributed to the analytical error displayed within laboratories (intralaboratory). There are many reasons for the interlaboratory variation that can be attributed to operational parameters such as mobile phase flowrate, mobile phase and buffer composition, vaporiser temperature, tip temperature and source temperature. 

Intralaboratory or within-laboratory precision refers to the precision of a test method when the results are obtained by the same operator in the same laboratory using the same apparatus. In some cases, the precision is applied to data gathered by a different operator in the same laboratory using the same apparatus. Thus, intralaboratory precision has an expanded meaning insofar as it can be applied to laboratory precision. 

The participants reported high precision of results (intralaboratory repeatability of measurements). The and M data obtained in particular laboratories scattered less than 3%, often even in the range of 1%. This is an excellent result, indeed. Unfortunately, the high repeatability of measurements may lead to a notion that the results are also exact. 

The infralaboratory calibration study was performed by the Institute for Environmental Studies. Sediment was extracted and cleaned up as indicated here. The determination of dioxin and/or dioxin-like content was according to the method indicated under the section DR CALUX analysis. For the intralaboratory study, the following parameters were investigated limit of detection (LOD), limit of quantitation (LOQ), and reproducibility and repeatability of the bioassay. 

The results of the multiple analysis of the standard 2,3,7,8-TCDD calibration curves are also be used to determine the per-participant LOD and LOQ taking into account interlaboratory variation (Table 3). This results show that on average, the partieipants of the calibration study meet the set LOD and LOQ derived from the intralaboratory study. 

Table 1 Intralaboratory repeatability and reproducibility of the dioxin response-chemically activated lucerferase (DR CALUX ) bioassay for sediment extracts 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD) TEQ = toxic equivalent.

TCDD calibration curves. From these data, a per-participant LQD and LQQ could be determined. Qn average, the participants of the calibration study met the set LQD and LQQ derived from the intralaboratory study. Furthermore, analysis of variance indicated that no significant differences in LOD between laboratories could be identified. 

For the determination of the intralaboratory repeatability of the DR CALUX bioassay for sediment samples, two sediment extracts were analyzed 10 times. Each analysis was performed in triplicate. As a prerequisite for a correct triplicate analysis, the percentage standard deviation in the triplicate determination should be below 15%. This is in accordance with the harmonized quality criteria for eell-based bioassay analyses of PCDDs/PCDFs in feed and food as formulated by Behnisch et al. (Behnisch et al, 2001 a) and as detailed in European Union directive 2002/69/ EC and direetive 2002/70/EC. The repeatability for the low-2,3,7,8-TCDD-content sediment... 

As with the determination of the intralaboratory repeatability, the intralaboratory reproducibility was determined by analyzing a cleaned sediment extract and a 3-pM 2,3,7,8TCDD standard on 10 separate days and by multiple persons. The reproducibility for the 3-pM 2,3,7,8-TCDD standard was found to be 13.8%, whereas the reproducibility for the cleaned sediment extraet was shown to be 19.9%. Since the observed reproducibilities are in the range of relative standard deviations for two sediment extracts analyzed in 10-fold on the same day (intralaboratory repeatability), the DR CALUX bioassay can be evaluated as a stable and robust bioanalytical tool. 

The in vitro bioassay for dioxins with cleaned sediment extracts (DR-CALUX) proved to comply with the QA/QC criteria needed to guarantee the reliability of data in an inter- and intralaboratory study (Besselink et al., 2004). The chemical stability of dioxins makes it possible to apply destructive clean-up procedures which remove all matrix factors. Sample extraction and cleanup for other in vitro bioassays for specific mechanisms of toxicity require further development to make sure that the chemicals of interest are not lost or unwanted chemicals included in the sediment extract to be tested. Table 4 summarizes possible bioassays that could be performed in addition to chemical analyses with the dredged sediment in a licensing system. 

Agencies or authorities such as ISO or lUPAC still do not provide any definition of ruggedness. In the chemical literature however, a ruggedness test was defined as [4,12] An intralaboratory experimental study in which the influence of small changes in the operating or environmental conditions on measured or calculated responses is evaluated. The changes introduced reflect the changes that can occur when a method is transferred between different laboratories, different experimentators, different devices, etc. . 

The run error is seen as a systematic error for one run and as a random variation over several runs performed intralaboratory. 

Validation-based approach Based on inter- or intralaboratory validation studies (precision, trueness, robustness) An extension of validation work, no extra work needed Some sources of uncertainly may be overlooked Eurachem [14], Barwick and Ellison [47]... 

Robustness-based approach Based on robustness tests as intralaboratory simulations of interlaboratory studies Simple, time-efficient approach Some sources of uncertainly may be overlooked method must first show to be robust Hund et al. [39]... 

Precision measures are divided into (1) repeatability precision measures s or SD (sT or SDr) and RSD (RSDr), (2) intralaboratory reproducibility precision or intermediate-precision measures SD and RSD, and (3) interlaboratory reproducibility precision s or SD (sR or SDR) and RSD (RSDr) [66]. 

The (intralaboratory tested) behavior of analytical process when small changes in environmental and/or operating conditions are made (generally used term)... 

Any or all of these conditions can be varied. To provide some guidance, intralaboratory reproducibility is used to express changes only within a laboratory, and interlaboratory reproducibility" is used to refer to the changes that occur between laboratories, for example in proficiency testing, interlaboratory method validation studies, and the like. Interlaboratory reproducibility is usually two to three times the repeatability. 

Figure 4.22. Pareto chart of the contributions to the uncertainty of the quantitative NMR analysis of Profenofos. The effects are a (intra), the intralaboratory precision P(std), the purity of the proton standard w, weighings of unknown and standard MW, the molecular weights of unknown and standard. (Data kindly supplied by T Saed Al-Deen.)...

The repeatability (sr) can be used to check duplicate repeats during normal operation of a method (see chapter 1). On its own, repeatability is not a complete basis for estimation of measurement uncertainty because it omits many effects that contribute to the bias of measurements made within a single laboratory. However, combined with a good estimate of the run bias, the intralaboratory precision, obtained from quality control data, can be used to give an estimate of measurement uncertainty. See section 6.6.3.2 for details on correction for bias and recovery. 

Metabolite Mean concentration Mean recovery (%) Mean intralaboratory imprecision (%) Mean interlaboratory imprecision (%)... 

Mitchell. A.D., Rudd, C.J. Caspary, W.J. (1988) Evaluation of the L5178Y mouse lymphoma cell mutagenesis assay intralaboratory results for sixty-three coded chemicals tested at SRI International. Environ, mol. Mutag., 12 (Suppl. 13). 37-101... 

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