4D analysis of lymphocyte movements in fresh lymph node tissue as a basis for future diagnostics in malignant lymphomas

Introduction While the clinical, morphological, genomic characteristics of lymphocytes in lymph node tissue are well known, little is known about the motility and interaction of primary human lymphocytes in lymph nodes. Therefore, the aim of this study was to demonstrate the movement of lymphocytes in fresh human lymph node tissue.

Material Methods Suspicious lymph nodes were exstirpated, representative tissue was submitted to routine diagnostic procedures, and a 2-mm piece of the remaining tissue was embedded in 5% low-gelling agarose. All subsequent processing and examination steps were carried out under special precautions to ensure survival and motility of the lymphocytes was not compromised. Slices 350 µm thick were cut and stained with antibodies (PD-1 and CD 20). These preparations were then supplied with oxygen, CO2 and nutrition in a special chamber and examined with a confocal laser microscope. For the four-dimensional analysis of cell migration, image stacks of 10-12 sections (z-step=5 µm) at depths of up to 80 µm were acquired every 10-20 s over a period of 20 min. The movements of the lymphocytes were automatically tracked and analysed using Imaris software.

Results Five cases of lymphadenitis (2 x EBV, 2 x non-specific and 1 x Kikuchi-Fujimoto) were studied. The mean velocities of the CD20-positive B cells was 5.73 µm/min. The mean velocity of PD-1 cells was 8.31 µm/min.

Discussion We succeeded in tracking the movement of lymphocytes in fresh human lymph node tissue and evaluating their velocity. This 4d approach to diagnostics will be increasingly important in the understanding and diagnosis of lymphomas in the future.

Publication History

Article published online:
24 May 2022