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A. Yoshida, H. Kawashima, T. Kaburaki, J. Hori, J. Numaga, Y. Fujino; Direct Confirmation of Inoculated Antigen Associated with Apc in the Spleen During Acaid Induction . Invest. Ophthalmol. Vis. Sci. 2003;44(13):755.
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Purpose: Anterior chamber-associated immune deviation (ACAID) induction appears to require an inoculated antigen’s entry, in association with APC, into spleen (SP). However, thus far, there has only been indirect evidence of this speculation and no direct proof. We, therefore, intend to detect this inoculated antigen in SP and cervical lymph node (CLN) both qualitatively and quantatively with reference to ACAID induction. Methods: Splenocytes (2x106) of enhanced green fluorescence protein (EGFP) mouse (C57BL/6) were injected, as antigen, into the anterior chamber (AC) or subconjunctival space (SS) of normal BALB/c mouse. One week after the injection, splenocytes (10x106) were injected subcutaneously. Another week after the subcutaneous injection, splenocytes (2x106) were injected into the auricle and the delayed-type hypersensitivity was measured to monitor ACAID induction. In separate experiments, SP and CLN of recipient mice were collected 1,3,6,12, and 24 hours after either AC or SS injection. Utilizing a confocal microscope, EGFP-positive cells were detected qualitatively and quantatively in these organs. Localization of macrophages (F4/80-positive cells) were also analyzed concurrently. Results: When splenocytes were injected into AC, ACAID was successfully induced in the recipient mice (AC-mice). However, ACAID was not induced in the recipient mice when injected into SS (SS-mice). As early as 3 hours after the injection into either AC or SS, EGFP-fluorescence was detected in association with F4/80-positive cells around the marginal sinus of both SP and CLN. Although there was little difference in the amount of fluorescence in SP, the amount of fluorescence in CLN of the SS-mice was significantly more than that of the AC-mice. Conclusions: As early as 3 hours after the injection, we could successfully detect injected antigen’s entry into SP in association with F4/80-positive macrophages after the injection to AC and SS. It appeared, though, that the amount of antigen in CLN of AC-mice was much less than that of SS-mice.
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