We investigated the clotting associated with delayed hypersensitivity (DH) responses in the mouse by sensitizing the animals to the contactant oxazolone (Ox), and then administering 125I-guinea pig fibrinogen i.v. 10 to 30 min before antigen challenge 5 days later. Early (4 to 8 hr) contact sensitivity (CS) responses in immunized mice were barely detectable by three conventional measures of CS, but the total 125I-cpm in ears challenged with hapten was 3.6 to 4.5 X that in control ears challenged with vehicle alone; moreover, the amount of urea-insoluble cpm (cross-linked 125I-fibrin-associated cpm) in the reactions to Ox was 6.5-fold to 8.2-fold that present in the control reactions. In 24 hr reactions that were near peak intensity by measurements of ear swelling, ear weight ratios, and ratios of 125I-5-iodo-2-deoxyuridine-labeled leukocyte infiltration, the cpm in antigen-challenged ears exceeded that in control ears by 13-fold to 53-fold. In addition, antigen-challenged ears contained 27 to 300 X the urea-insoluble cpm present in control ears. 125I-Fibrin deposition was not a specific characteristic of CS reactions, because a small amount of urea-insoluble reactivity was also detected in some reactions to Ox in native mice. Nevertheless, the assay was exquisitely sensitive and readily detected quantitative differences between the immunologically specific and nonspecific reactions at very early intervals after challenge or with suboptimal doses of antigen. Furthermore, it was more sensitive than conventional tests of CS in detecting the reactions elicited in mice that had been passively sensitized to Ox by adoptive transfer of immune lymph node cells. Finally, we showed that the assay gave similar results when we tested CS reactions elicited in mast cell deficient WBB6F1-W/Wv and littermate normal (+/+) mice, demonstrating yet another similarity in the phenotype of DH reactions elicited in the presence or absence of mast cells.

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