Protocol sample size

Clinical trials are costly to conduct, and results are often critical to the commercial viability of a phytochemical product. Seemingly minor decisions, such as which measurement tool to use or a single entry criterion, can produce thousands of dollars in additional costs. Likewise, a great deal of time, effort and money can be saved by having experts review the study protocol to provide feedback regarding ways to improve efficiency, reduce subject burden and insure that the objectives are being met in the most scientifically sound and cost-effective manner possible. In particular, I recommend that an expert statistician is consulted regarding sample size and power and that the assumptions used in these calculations are reviewed carefully with one or more clinicians. It is not uncommon to see two studies with very similar objectives, which vary by two-fold in the number of subjects under study. Often this can be explained by differences in the assumptions employed in the sample size calculations. 

The first step in all RNA isolation protocols involves lysing the cell in a chemical environment that denatures ribonucleases. The RNA is then fractionated from other cellular macromolecules by either homogenizing the tissue (dissected brain tissue) or simply vortexing the sample (very small tissues and laser-microdissected sample) without further homogenization. The cell type from which the RNA is to be isolated, the sample size, and the eventual use of the RNA will determine which procedure described here is appropriate. 

Results on the refinement of protocols for sampling analysis are forthcoming in the report on the EPA/NBS workshop on monitoring. Additional work on statistical design is also in progress to find practical ways to reduce required sample size which in turn leads to reductions in cost and more efficiency in dealing with contractor release after abatement. 

Clinical trials generate vast quantities of data, most of which are processed by the sponsor. Assessments should be kept to the minimum that is compatible with the safety and comfort of the subject. Highest priority needs to be given to assessment and recording of primary endpoints, as these will determine the main outcome of the study. The power calculation for sample size should be based on the primary critical endpoint. Quite frequently, trials have two or more evaluable endpoints. It must be stated clearly in the protocol whether the secondary endpoints are to be statistically evaluated, in which case power statements will need to be given, or are simply... 

Does basing the sample size calculation on the per-protocol set and then increasing the sample size to allow for dropouts ensure that the per-protocol set will not be subject to bias ... 

No It often makes sense to power for the per-protocol set and then factor upwards to allow for dropouts as this will also ensure that there is enough power for the full analysis set providing any extra patient-to-patient variation in the full analysis set does not counterbalance the increase in sample size, but the analysis based on the per-protocol set is still subject to bias. See Section 8.5.2 for further discussion on this point. 

Generally speaking we power based on the per-protocol set and then increase the sample size requirement to give the number required in the full analysis set. Under some circumstances, for example in anti-infective trials, we factor up further to take into account the patients that are recruited, but are not eligible for the full analysis set. 

It must be noted, however, that even if the sample size calculation gives enough power for the per-protocol analysis the potential for bias in that analysis still remains. 

A detailed statement of the basis of the sample size calculation should be included in the protocol and in the final report. This statement should contain the following ... 

There may, of course, be cases, especially in the early exploratory phase, where the sample size has been chosen on purely practical or feasibility grounds. This is perfectly acceptable in that context and the sample size section in the protocol should clearly state that this is the case. 

As with sample size in superiority trials we generally power on the basis of the per-protocol set and increase the sample size to account for the non-evaluable patients. This is particularly important in non-inferiority trials where the full analysis set and the per-protocol set are co-primary analyses. Note also, as before in superiority trials further factoring up may be needed if there are randomised patients who are being systematically excluded from the full analysis set, as is the case, for example, in anti-infective trials. 

A revised sample size may then be calculated using suitably modified assumptions, and should be justified and documented in a protocol amendment and in the clinical study report... The potential need for re-estimation of the sample size should be envisaged in the protocol whenever possible. ... 

The clinical endpoint is a clinically meaningful measure of how patients feel, function or survive. Investigator-rated or self-assessed rating instruments are the most frequently used clinical endpoints. A primary endpoint is the main outcome that a study protocol is designed to evaluate. The statistical power and the sample size calculation of a particular trial are determined by the primary endpoint. Depending on the purpose of a study the primary endpoint can be... 

Other than the time specified in the individual steps of each protocol, the total time spent on each methylation procedure depends largely on the sample size, the time required for evaporation of organic solvents, and the availability of space for sample tubes in the heating block and the manifold of the nitrogen-stream evaporator. 

As the levels of tocopherols and tocotrienols in meat samples are usually lower than in oil and fat samples, a larger sample size is needed in the sample preparation. The meat sample is homogenized to weaken the sample matrix. As in Basic Protocol 2, saponification, heating, and liquid/liquid extraction are used to increase the recovery and remove interference compounds. Satisfactory results can be achieved using a reversed-phase HPLC method. 

Because the objective of the protocol was not only to get a quantitative elemental analysis, but also to keep the sample size as small as possible, EDS must be considered as preliminary test for elemental analysis by ICP-MS or ICP-OES, thus allowing for the chemical composition of die material to dictate what must be done next... 

M NaCl, pH 7.2, containing 10 mM EDTA as the buffer. To obtain efficient separation between the reduced antibody and excess reductant, the sample size applied to the column should be at a ratio of no more than 5% sample volume to the total column volume. Collect 0.5-ml fractions and monitor for protein at 280 nm. To monitor the separation of the second peak (excess Traut s reagent), the BCA protein assay reagent (Pierce) may be used according to the procedure described in the previous section, protocol step 4. 

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