Fluorine-18 labeled mouse bone marrow-derived dendritic cells can be detected in vivo by high resolution projection imaging
Introduction
Active immunotherapy using tumor antigen loaded DC in mice have been extensively studied Celluzzi et al., 1996, Porgador et al., 1996, Zitvogel et al., 1996, Lotze et al., 1997, Lotze et al., 2000, Tuting et al., 1997, Fields et al., 1998a, Fields et al., 1998b, Nestle et al., 1998, Salgaller et al., 1998. Other studies demonstrated protection against viruses (De Bruijn et al., 1998) and bacteria (Mbow et al., 1997). Furthermore, human trials utilizing DC loaded with antigen are underway in several institutions. The induction of immunity depends on the interaction between DC and T cells. However, the best route of DC administration for ensuring migration to the T cell areas of lymph nodes, thereby ensuring optimal interactions, is uncertain, in part, because the sites where human DCs localize after injection are not fully known. Although bone marrow-derived DCs (BMDC) or their precursors circulate in the peripheral blood and subsequently reside in peripheral tissues, acquire antigen and migrate to regional lymph nodes in physiological circumstances [Steinman, 1991 #63; Banchereau, 1998 #62; Flores-Romo, 2001 #1], the migration pattern is still under intensive investigation. Migration of dendritic cells after intravenous, intradermal or subcutaneous injection has been assessed by using different tracing methods, such as membrane or cytosolic fluorescent dyes Austyn et al., 1988, Barratt-Boyes et al., 1997, Ingulli et al., 1997, Lappin et al., 1999 or cells labeled with radionuclides such as Cr-51 Cruz et al., 1990, Saeki et al., 1999 or Indium-111 Kupiec-Weglinski et al., 1988, Larsen et al., 1990, Morse et al., 1999, Thomas et al., 1999. In most of these studies the organs are removed at various times after injection of the label.
The aim of our study was to develop a method to label DC with the positron emitting radionuclide F-18, in order to detect DC migration in vivo by a high-resolution small animal PET camera. Here we describe our labeling method, the effects of labeling on the viability, phenotype and function of BMDC in vitro, and how F-18 labeled cells can be visualized and followed by a high resolution animal scanner.
Section snippets
Mice
Female BALB/c (I-Ad), C57BL/6 (I-Ab) and F1 (BALB/cxC57BL/6) mice (6–10 weeks old) were obtained from the National Institutes of Health animal facility. All animals were treated in accordance with institutional protocol.
Cells
BMDC were generated using the method of Fields et al. (1998a) with minor modifications. Briefly, bone marrow was flushed from tibias and femurs of female BALB/c or C57BL/6 mice and depleted of red blood cells with ammonium chloride. Cells were plated at 1×106 cell/ml in T162 T
Labeling efficiency and retention of radionuclide with BMDC
The F-18 labeling efficiency—percentage of original activity used for labeling remained with the cells after labeling and three washes—was 19±4% (n=18). Based on the specific activity of F-18, the cells were labeled with an average of 107 label/cell. Fig. 1 shows the distribution of F-18 in cells and supernatants as a function of time after labeling. At 37 °C considerable release occurred during the first hour and then remained essentially unchanged. The release of F-18 by BMDC may result from
Discussion
This is the first description of a method for labeling mouse BMDC with a positron emitting radionuclide F-18 in order to follow their migration with a high-resolution imager in vivo. F-18 is a radionuclide with a short (109.7 min) half life, and a high specific activity (>200 Ci/mmol at EOB). The short half life allows detection of the radionuclide for a maximum of 5–6 h, which in the case of BMDC migration is rather short, since the maximum number of injected BMDC can be found in the draining
Acknowledgements
The authors would like to thank to Elaine Jagoda, John L. Holt for helpful discussions and Jay Linton for providing expert technical help.
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