Application of a modified laser diffractometry method for assessing the morphological parameters of human and rat erythrocytes

Abstract

Size characteristics of erythrocytes (MCV and RDW) are among the parameters that have diagnostic value in establishing various diagnoses. In clinical laboratories, these parameters are determined using cytometers, however, there is interest in developing other methods that allow rapid acquisition of quantitative information from large cell ensembles. The main focus of this work is on measuring the mean size (diameter) of erythrocytes, which can be related to the MCV parameter [1], by the method of laser diffractometry. Although the capabilities of this method are not limited to calculating the mean cell diameter, its correct measurement, as well as comparison with data from the reference method, microscopy, is a necessary step in developing the methodology for conducting laser diffractometry experiments.
This study investigates wet smears [2] with blood samples from healthy human donors (32 smears, 6 donors) and laboratory rats (12 smears, 4 rats; ethical approval No. 183-д-з dated 18.06.2026) using laser diffractometry and microscopy. It was shown that a unified experimental protocol for sample preparation and measurement is applicable to all blood samples.
During the experiments, it was observed that, on average, the diameters obtained by microscopy were larger than those calculated by laser diffractometry. To relate the two methods, an empirical calibration correction, termed the “instrument factor,” was introduced. The instrument factor is defined as the ratio of the erythrocyte diameter in a smear calculated by microscopy to the diameter calculated by diffractometry. This ratio is determined for all studied samples, then the mean value is computed and taken as the instrument factor for the entire sample set.
Taking into account that the ratio between the diameters calculated by the two methods is on average close to 1.03 for humans and 1.27 for rats, and using these values as the instrument factors, the difference between the two methods within the proposed approach is 2.2% and 3.7% for human and rat blood samples, respectively. The reasons for this difference require separate investigation. Nevertheless, for quantitative interpretation of diffraction patterns, it is necessary to account for the specific features of the particular cell ensemble, and the instrument factor should be determined separately for each type of sample.

The study was conducted with the financial support of the Russian Science Foundation (grant No 26-15-00086). M.S. Lebedeva is a scholarship holder of the "BASIS" foundation (grant No 23-2-2-36-1).

1. Дубровский В.А., Торбин С.О., Забенков И.В. Определение индивидуальных и средних характеристик эритроцитов нативной крови методом статической спектральной цифровой микроскопии // Оптика и спектроскопия. 2022. Т. 130. No 6. С. 894–905.
2. Лебедева М.С., Цыбров Е.Г., Никандрова А.Е. Эксперименты по лазерной дифрактометрии мазков крови // Оптические методы исследования потоков: труды XVI Международной научно-технической конференции, 28 июня – 02 июля 2021. М.: Издательство «Перо», 2021. С. 175–182.
https://omfi-conf.ru/omfi2021/OMFI-2021-Proceedings.pdf


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Maria S. Lebedeva
Lomonosov Moscow State University
Россия

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