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

Background: 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. Objective: We set out to determine the amount of time that specimens can remain in 10% buffered formalin and still retain their diagnostic properties. Methods: 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. Results: 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. Limitations: Sample size, only examining 3 disease processes, and samples already having been in Michel medium were the major limitations in the study. Conclusion: 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.