ABSTRACT
Working with infectious agents that require BSL-3 level containment agents offers many challenges for researchers. BSL-3 containment laboratories are usually not equipped with expensive specialty equipment that is needed for studies such as flow cytometric analysis, microscopy, and proteomic analyses. Therefore, for most researchers that are working with BSL-3 level infectious agents, removal of samples from BSL-3 labs for these types of studies is necessary, and methods for complete and dependable inactivation of the samples are required. In this report we have done a thorough characterization of the effectiveness of paraformaldehyde fixation for inactivation of cell samples infected with the intracellular bacterial agents Burkholderia pseudomallei (Bp) and Francisella tularensis (Ft), both of which are Tier 1 select agent pathogens that require BSL-3 containment. We have demonstrated that cells infected with these pathogens are completely inactivated via 5-minute treatment with 4% paraformaldehyde. Moreover, a 15-minute treatment with 2% paraformaldehyde completely sterilized both Bp- and Ft-infected cells. These studies …show more content…
Due to the expense of this type of equipment and the technical expertise required for proper operation, flow cytometers/cell sorters are typically found in shared core facilities that employ dedicated operators for analysis of a variety of sample types. This creates many challenges for researchers working with agents requiring biosafety level 3 (BSL-3) containment, since most core flow cytometry facilities are located in laboratories with biosafety level 2 (BSL-2) containment capabilities. Additional factors, such as biosecurity, play a role when the agents in use are considered select agents and these security procedures must be maintained until the samples are rendered
Working with infectious agents that require BSL-3 level containment agents offers many challenges for researchers. BSL-3 containment laboratories are usually not equipped with expensive specialty equipment that is needed for studies such as flow cytometric analysis, microscopy, and proteomic analyses. Therefore, for most researchers that are working with BSL-3 level infectious agents, removal of samples from BSL-3 labs for these types of studies is necessary, and methods for complete and dependable inactivation of the samples are required. In this report we have done a thorough characterization of the effectiveness of paraformaldehyde fixation for inactivation of cell samples infected with the intracellular bacterial agents Burkholderia pseudomallei (Bp) and Francisella tularensis (Ft), both of which are Tier 1 select agent pathogens that require BSL-3 containment. We have demonstrated that cells infected with these pathogens are completely inactivated via 5-minute treatment with 4% paraformaldehyde. Moreover, a 15-minute treatment with 2% paraformaldehyde completely sterilized both Bp- and Ft-infected cells. These studies …show more content…
Due to the expense of this type of equipment and the technical expertise required for proper operation, flow cytometers/cell sorters are typically found in shared core facilities that employ dedicated operators for analysis of a variety of sample types. This creates many challenges for researchers working with agents requiring biosafety level 3 (BSL-3) containment, since most core flow cytometry facilities are located in laboratories with biosafety level 2 (BSL-2) containment capabilities. Additional factors, such as biosecurity, play a role when the agents in use are considered select agents and these security procedures must be maintained until the samples are rendered