Dye amount quantification of Papanicolaou-stained cytological images by multispectral unmixing: spectral analysis of cytoplasmic mucin.

Purpose: The color of Papanicolaou-stained specimens is a crucial feature in cytology diagnosis. However, the quantification of color using digital images is challenging due to the variations in the staining process and characteristics of imaging equipment. The dye amount estimation of stained specimens is helpful for quantitatively interpreting the color based on a physical model. It has been realized with color unmixing and applied to staining with three or fewer dyes. Nevertheless, the Papanicolaou stain comprises five dyes. Thus, we employ multispectral imaging with more channels for quantitative analysis of the Papanicolaou-stained cervical cytology samples. Approach: We estimate the dye amount map from a 14-band multispectral observation capturing a Papanicolaou-stained specimen using the actual measured spectral characteristics of the single-stained samples. The estimated dye amount maps were employed for the quantitative interpretation of the color of cytoplasmic mucin of lobular endocervical glandular hyperplasia (LEGH) and normal endocervical (EC) cells in a uterine cervical lesion. Results: We demonstrated the dye amount estimation performance of the proposed method using single-stain images and Papanicolaou-stain images. Moreover, the yellowish color in the LEGH cells is found to be interpreted with more orange G (OG) and less Eosin Y (EY) dye amounts. We also elucidated that LEGH and EC cells could be classified using linear classifiers from the dye amount. Conclusions: Multispectral imaging enables the quantitative analysis of dye amount maps of Papanicolaou-stained cytology specimens. The effectiveness is demonstrated in interpreting and classifying the cytoplasmic mucin of EC and LEGH cells in cervical cytology.