The effect of reducing the number of cells scored on the performance of the in vivo rat liver UDS assay.
The most labour-intensive feature of the in vivo rat liver UDS assay is the scoring of hepatocyte autoradiograms by microscope. Even with image analyser and computer equipment the scoring phase of a full study might require half of the technical effort applied. Practice recommended by guidelines has been to score 50 cells/slide and two slides per animal. Now sufficient data have been accumulated, an evaluation was made to observe whether this procedure was necessary. An analysis of the accumulated UDS database in our laboratory was made to determine the sources of variability of mean net nuclear grain count, [N-C]. It was observed that the two largest components of variation in negative control animal mean [N-C] were between-day and interanimal variability. The contribution from sampling error during slide scoring was relatively small. Theoretical calculations showed that the greater sampling error derived from scoring 30 rather than 50 cells/slide would result in only a marginal increase in total assay variation. To test this, 30 cells/slide were randomly selected from the 50 cells scored originally in negative control animals in each of 18 studies over an 18-month period. It was confirmed that reducing the number of cells had a negligible effect on the variation of negative control animal mean [N-C] values. Furthermore, analysis of data from 10 more studies confirmed that within-study variation would be essentially unaffected by scoring 30 cells/slide. The use of 30 rather than 50 cells per slide (a total of 60 cells per animal) has therefore been adopted for all current studies and scoring procedures modified to avoid operator bias during the selection of a smaller number of cells.
Kennelly
J C
JC
Zeneca Central Toxicology Laboratory, Macclesfield, Cheshire, UK.
Pate
I
I
Greenwood
M R
MR
D000818:Animals; D002452:Cell Count; D002467:Cell Nucleus; D003627:Data Interpretation, Statistical; D007091:Image Processing, Computer-Assisted; D008099:Liver / Q000166:cytology / Q000187:drug effects*; D009152:Mutagenicity Tests / Q000379:methods*; D051381:Rats; D015983:Selection Bias