Two-Photon Imaging of Lymphocyte Motility and Antigen Response in Intact Lymph Node

Abstract

Lymphocyte motility is vital for trafficking within lymphoid organs and for initiating contact with antigen-presenting cells. Visualization of these processes has previously been limited to in vitro systems. We describe the use of two-photon laser microscopy to image the dynamic behavior of individual living lymphocytes deep within intact lymph nodes. In their native environment, T cells achieved peak velocities of more than 25 micrometers per minute, displaying a motility coefficient that is five to six times that of B cells. Antigenic challenge changed T cell trajectories from random walks to "swarms" and stable clusters. Real-time two-photon imaging reveals lymphocyte behaviors that are fundamental to the initiation of the immune response.

Visualizing dendritic cell migration within the skin

“…Note that the resolution of individual ECM Wbers is superior in the forward direction. Scale bars 20 m Since interstitial leukocyte migration within tissue is dependent on temperature (Miller et al 2002), the temperature of the ear platform is regulated independently and maintained at 36°C, while the body temperature is kept at 37°C through a heating pad underneath the mouse. Body temperature is monitored through a rectal probe.…”

T cell–antigen-presenting cell interactions visualized in vivo in a model of antigen-specific inflammation

“…The absence of this specialized matrix in the iris stroma likely contributes to the slower speed that we observed for T cells migrating in the iris. Estimates of the average lateral speed of T cells in lymph node vary from 4 µm/min during an activation phase to about 11 µm/min in the absence of an antigenic challenge [4,6,33]. These are considerably faster than the 1-2 µm/min lateral speed reported here and likely reflect tissue-specific differences in the nature of chemotactic signals and/or the extracellular matrix along which the cells must navigate.…”

Sculpting the immunological response to dengue fever by polytopic vaccination

“…This process leads to 0.5% diversity of the memory repertoire, because 0.58 10 ≈ 0.5%. In vivo T cell tracking experiments have shown that T cells continue to interact with antigen in the lymph node up to 5 days after initial stimulation [41,42], and examinations of draining lymph nodes have found that antigen capable of priming naive T cells is present up to 7 days post inoculation [43], both of which support the concept that the selection process may continue during each T cell division. Although the exact mechanism of the T cell expansion during the primary immune response remains elusive, it is observed that the expansion of the T cells is nonlinear [44], and that in most cases there is competition among the T cells for the presented antigen [45,46].…”