Reliability of Nucleus-To-Cell and Nucleus-To-Cytoplasm Calculations for Conjunctival Impression Cytology Specimens

Abstract

Purpose: To assess the outcome of two different nucleo-cytoplasmic ratio calculations in relation to the number of cells measured from conjunctival impression cytology (CIC) images.

Methods: CIC from the exposed nasal bulbar conjunctiva surface was undertaken on 40 young adults using a Millcell^®-CM filter; the filters were stained with Giemsa, colour images taken of the cells in a monolayer at 200 X magnification, graded by the Nelson scale and then a projection overlay method used to outline the cell and nucleus borders. Nucleo-cytoplasmic ratios were calculated based on literal interpretation of the term (i.e. N/CYT = nucleus area/[cell area − nucleus area]) as well as based on cell area (i.e. N/CELL = nucleus area/cell area).

Results: A range of N/CYT or N/CELL values are expected for all cells, regardless of squamous metaplasia grade, with slightly greater heterogeneity expected from N/CYT calculations. The difference between the two calculations gets progressively less as the squamous metaplasia grade increases. For all grades of cells, these ratios could be determined to within approximately ±1% if adequate numbers of cells were measured (i.e. 75, 65, 50 and 35 for grade 0 to grade 3 cells respectively). If fewer cells are measured, then the variability in N/CELL or N/CYT calculations can be expected to increase.

Conclusions: In assessments of nucleo-cytoplasmic ratios, it is important that the calculation used be specified and that the numbers of cells measured be provided.

Keywords::

• Impression cytology
• Bulbar conjunctiva
• Human
• Squamous metaplasia
• N/C ratio

Acknowledgements

The assistance of Dr. Taher Naase, Alexia Bibollet and Louise Gess in the collection of some of the samples used in these analyses is gratefully acknowledged.

Declaration of interest: The author has no proprietary interest in any of the products or procedures outlined in this paper.