Cytokine measurement in biological samples after physicochemical treatment for inactivation of biosafety level 4 viral agents

Abstract

Physicochemical techniques such as gamma-irradiation, membrane disruption by detergents like sodium dodecyl sulfate (SDS), and fixation with formaldehyde or paraformaldehyde are routinely used to inactivate biological specimens from patients and animals infected with Filoviruses and Arena viruses that must be studied in BSL 4 facilities. The effects of these inactivation techniques on the levels of immunologically active proteins like cytokines and chemokines are not known. Therefore, we investigated the effect of several decontamination techniques on the immunoreactivity and bioactivity of the inflammatory cytokines IL-1beta, IL-6, TNF-alpha, IL-12, and IFN-gamma, the anti-inflammatory cytokine IL-10, and the chemokine IL-8 in biological specimens. SDS (96%-100% reduction), paraformaldehyde treatment (11%-100% reduction), and heat denaturation (75%-100% reduction) were found to decrease markedly the levels of all cytokines and chemokines as measured by enzyme-linked immunosorbent assays. In contrast, gamma-irradiation was found to have little or no effect on the immunoreactivity of these cytokines/chemokines and on the biological activity of tumor necrosis factor (TNF) alpha. Our data suggest that, of the agents tested, gamma-irradiation is the preferred technique for inactivation of biological specimens containing viral agents that require the use of BSL 4 for immunological studies.