Label-free hematology analysis method based on defocusing phase-contrast imaging under illumination of 415 nm light.

Label-free imaging technology is a trending way to simplify and improve conventional hematology analysis by bypassing lengthy and laborious staining procedures. However, the existing methods do not well balance system complexity, data acquisition efficiency, and data analysis accuracy, which severely impedes their clinical translation. Here, we propose defocusing phase-contrast imaging under the illumination of 415 nm light to realize label-free hematology analysis. We have verified that the subcellular morphology of blood components can be visualized without complex staining due to the factor that defocusing can convert the second-order derivative distribution of samples' optical phase into intensity and the illumination of 415 nm light can significantly enhance the contrast. It is demonstrated that the defocusing phase-contrast images for the five leucocyte subtypes can be automatically discriminated by a trained deep-learning program with high accuracy (the mean F1 score: 0.986 and mean average precision: 0.980). Since this technique is based on a regular microscope, it simultaneously realizes low system complexity and high data acquisition efficiency with remarkable quantitative analysis ability. It supplies a label-free, reliable, easy-to-use, fast approach to simplifying and reforming the conventional way of hematology analysis.