Which is NOT relevant to a laboratory diagnosis of smallpox variola?

Direct fluorescent antibody (DFA) test

DFA staining of clinical specimens directly on a slide (e.g., smears from skin lesions) has proved to be a rapid, sensitive, and specific method that can be performed in a BSL-3 laboratory. DFA was the mainstay of laboratory diagnostics for smallpox in a reference laboratory of WHO.

Electron microscopy

Negative-contrast EM allows rapid laboratory identification of smallpox virions by their characteristic morphology, which is distinct from the appearance of other poxviruses. Smallpox virus particles seen using EM in clinical samples of skin lesions are typically spherical, 250–270 nm in diameter, and contain the typical "dumbbell-shaped" lateral bodies.

Nucleic acid amplification tests (NAATs)

The PCR assay targeting a region of the A-type inclusion (ATI) gene proved to be a very sensitive laboratory test that allowed reliable detection and differentiation of smallpox from other orthopoxviruses. Primers and probes selected from conserved regions of the ATI gene have been successfully tested for detection of variola virus DNA in clinical smallpox specimens.

Histopathology

Light microscopic examination of skin lesion histopathology can provide useful diagnostic clues if there is a high index of suspicion of smallpox, especially if the tissue is collected early (4–6 days) in the evolution of the lesions. The main features of smallpox skin lesion histology are (a) epidermal hyperplasia with ballooning degeneration, necrosis, and formation of multinucleated giant cells; (b) dermal edema associated with vasculitis and a perivascular infiltrate; and (c) presence of intracytoplasmic inclusion bodies in the epidermis.