Rectal epithelial cell proliferation: Comparison of errors of measurement with inter-subject variance

If measures of rectal epithelial kinetics are to be used on a large scale, or as part of multicenter studies, the variance of repeated measures must be sufficiently small to detect the effect of clinical status and dietary, or chemoprevention interventions. Errors in measurement can be categorized as intra-reader (σ²(I)), inter-biopsy (σ²(B)) inter-laboratory (σ²(L)), and inter-reader (σ²(R)); the sum of these (σ²(T)) When compared to inter-subject variance (σ²(S)) is a measure of the precision with which cell kinetics can be estimated for a given subject. We estimated the variances of the component errors in four separate experiments, each involving 6 to 10 subjects, using the proliferating cell nuclear antigen (PCNA) technique. PCNA labeling index variances for a single laboratory study were σ²(I) = 0.02-0.07 and σ²(B) = 0.08-0.15. Inter-laboratory and inter-reader variances were σ²(L) = 0.08 and σ²(R) = 0.03. Summing the errors applicable within a laboratory gives a rounded estimate of 0.15. For inter-laboratory studies the total error (σ²(T)) is greater, estimated in our two-laboratory study as 0.3. In a separate study, the variation between biopsies (σ²(B)) could be accounted for by the variation between crypts within biopsies; an acceptable level of accuracy is achieved after counting 20-30 crypts. In contrast to the measurement errors, the variance between subjects, which includes a treatment effect of diet, was larger, estimated to be between 0.75 and 1.30. Immunohistochemical staining of PCNA offers a reproducible estimate of cell kinetics, with the variance of the total error in measurement approximately one-quarter of the inter-subject variance. While further studies are needed to determine within-subject variance over time and its biological relationship to cancer risk, this method is sufficiently precise for larger-scale clinical studies.