A Radioisotopic Method to Measure Delayed Type Hypersensitivity in the Mouse

Abstract

Delayed type hypersensitivity (DTH) lesions have been difficult to evaluate objectively in the mouse because they are usually assessed in terms of an increase in footpad swelling or ear thickness. We have developed a radioisotopic method which not only reduces the observer’s error but also gives an objective measurement of the cellular activity in the lesions. In brief, 10 μl of the test antigen is injected intradermally into the left pinna and either nothing or the same volume of a control solution into the right. 10 h later, a 2-μCi pulse of 5-iodo-2′deoxyuridine-¹²⁵I is given intravenously, the ears cut off at the hairline 16 h later and the radioactivity counted in a gamma spectrometer. The following was obtained as evidence that the increased radioactivity of the left pinna over the right was a measure of the extent of a DTH response: (1) the ear reaction was delayed in mice without serum antibodies becoming maximum at 24 h; (2) there was a mononuclear cell infiltration in the left pinna and autoradiographs revealed radioactive label bound to these cells; (3) athymic mice could not develop a 24-hour ear reaction, and (4) antigens known not to activate T cells did not elicit the ear response. Cell transfer studies will be described in a subsequent paper. Different sensitization regimes were required with different antigens in order to obtain the highest levels of DTH as tested by the ear response, and the maximal ear reaction occurred at different days. The ear reaction showed the specificity expected of a DTH response.

H-2 gene complex restricts transfer of delayed-type hypersensitivity in mice.

“…The test used for DTH has been described in detail elsewhere (11,12). It depends on the labeling of replicating cells by '25I-dUrd and the quantification of the influx of labeled mononuclear cells (presumably monocytes) into a defined site where antigen is deposited as a challenge (left ear).…”

“…The present investigations were designed to determine whether successful transfer of DTH by lymphoid cells from sensitized to naive mice might be restricted by genes of the MHC. These studies were made possible by the use of congenic strains of mice and of a new radioisotopic assay which gives an objective measure of the extent of DTH in mice (11,12).MATERIALS AND METHODS Mice. Female mice (2-3 month old) were used for sensitization.…”

Application of a radiometric ear assay for studies of adjuvant arthritis in rats

“…For every antigen utilized in the radiometric ear assay with T U d R , the possibility of a nonspecific inflammatory response must be eliminated. Therefore, a dose-response curve was obtained to establish a challenge dose of M butyricum which produced an ear ratio less than 1.5 when injected into nonsensitized rats (10). Table 4 shows the results of such a challenge dose-range study in Lewis-Wistar rats suffering from adjuvant disease, and in nonsensitized counterparts.…”

“…Several more objective methods have been described for the measurement of delayed hypersensitivity (7-lo), and one technique that allows discrimination between cellular response and nonspecific inflammation under defined conditions is the method developed and characterized in the mouse by Vadas et a1 (10). This radioisotopic assay for determination of DTH in vivo using 5-iodo-2'-deoxyuridine-'z51 ('"IUdR) measures, in a sensitive and reproducible fashion, the influx of labeled mononuclear cells into DTH lesions in the ear.…”

“…This radioisotopic assay for determination of DTH in vivo using 5-iodo-2'-deoxyuridine-'z51 ('"IUdR) measures, in a sensitive and reproducible fashion, the influx of labeled mononuclear cells into DTH lesions in the ear. It has been used by Vadas et a1 (10) and others (11,12) with conventional sensitizing agents, e.g., sheep erythrocytes and 2,4-dinitrofluorobenzene, as well as with methylated-bovine serum albumin, an agent that produces an arthropathy in the mouse.…”