Can direct immunofluorescence testing still be accurate if performed on biopsy specimens after brief inadvertent immersion in formalin?

Abstract

Direct immunofluorescence is useful in the diagnosis of autoimmune, vesiculobullous, and connective tissue diseases. Michel medium is typically indicated for transport, but clinicians may inadvertently place samples into formalin. We set out to determine the amount of time that specimens can remain in 10% buffered formalin and still retain their diagnostic properties. Biopsy samples were examined from cases with established diagnoses of bullous pemphigoid (n = 12), dermatitis herpetiformis (n = 6), and pemphigus vulgaris (n = 6) and exposed to formalin for time points ranging from 2 minutes to 4 hours. We found that immunoreactants were detectable in the majority of samples when subjected to 2 minutes of formalin exposure. Dermatitis herpetiformis and pemphigoid samples retained immunogenicity for 10 minutes, whereas pemphigus showed reduced immunogenicity for all samples studied. A nonimmunologic nuclear fluorochroming pattern was noted in some of the specimens after formalin immersion. Sample size, only examining 3 disease processes, and samples already having been in Michel medium were the major limitations in the study. In direct immunofluorescence studies, formalin exposure to biopsy specimens causes two types of artifactual changes: (1) the shortest exposure (2 minutes) causes complete loss of diagnostic markers of pemphigus; and (2) prolonged exposure changes tissue to a form that allows fluorescein-labeled antibodies to give fluorochroming reactions of nuclei (which can be mistaken for in vivo antinuclear antibody reactions of lupus erythematosus). After time intervals of 10 minutes to 2 hours, direct immunofluorescence studies of proven cases of bullous pemphigoid and dermatitis herpetiformis retained variable levels of specific reactivity.