Rapid establishment of the European Bank for induced Pluripotent Stem Cells (EBiSC) - the Hot Start experience.

A fast track "Hot Start" process was implemented to launch the European Bank for Induced Pluripotent Stem Cells (EBiSC) to provide early release of a range of established control and disease linked human induced pluripotent stem cell (hiPSC) lines. Established practice amongst consortium members was surveyed to arrive at harmonised and publically accessible Standard Operations Procedures (SOPs) for tissue procurement, bio-sample tracking, iPSC expansion, cryopreservation, qualification and distribution to the research community. These were implemented to create a quality managed foundational collection of lines and associated data made available for distribution. Here we report on the successful outcome of this experience and work flow for banking and facilitating access to an otherwise disparate European resource, with lessons to benefit the international research community. ETOC: The report focuses on the EBiSC experience of rapidly establishing an operational capacity to procure, bank and distribute a foundational collection of established hiPSC lines. It validates the feasibility and defines the challenges of harnessing and integrating the capability and productivity of centres across Europe using commonly available resources currently in the field.

Check your cultures! A list of cross-contaminated or misidentified cell lines.

Continuous cell lines consist of cultured cells derived from a specific donor and tissue of origin that have acquired the ability to proliferate indefinitely. These cell lines are well-recognized models for the study of health and disease, particularly for cancer. However, there are cautions to be aware of when using continuous cell lines, including the possibility of contamination, in which a foreign cell line or microorganism is introduced without the handler's knowledge. Cross-contamination, in which the contaminant is another cell line, was first recognized in the 1950s but, disturbingly, remains a serious issue today. Many cell lines become cross-contaminated early, so that subsequent experimental work has been performed only on the contaminant, masquerading under a different name. What can be done in response-how can a researcher know if their own cell lines are cross-contaminated? Two practical responses are suggested here. First, it is important to check the literature, looking for previous work on cross-contamination. Some reports may be difficult to find and to make these more accessible, we have compiled a list of known cross-contaminated cell lines. The list currently contains 360 cell lines, drawn from 68 references. Most contaminants arise within the same species, with HeLa still the most frequently encountered (29%, 106/360) among human cell lines, but interspecies contaminants account for a small but substantial minority of cases (9%, 33/360). Second, even if there are no previous publications on cross-contamination for that cell line, it is essential to check the sample itself by performing authentication testing.

In vivo function of a gammaherpesvirus virion glycoprotein: influence on B-cell infection and mononucleosis.

The human gammaherpesviruses Epstein-Barr virus and Kaposi Sarcoma-associated herpesvirus both contain a glycoprotein (gp350/220 and K8.1, respectively) that mediates binding to target cells and has been studied in great detail in vitro. However, there is no direct information on the role that these glycoproteins play in pathogenesis in vivo. Infection of mice by murid herpesvirus 4 strain 68 (MHV-68) is an established animal model for gammaherpesvirus pathogenesis and expresses an analogous glycoprotein, gp150. To elucidate the in vivo function of gp150, a recombinant MHV-68 deficient in gp150 production was generated (vgp150Delta). The productive viral replication in vitro and in vivo was largely unaffected by mutation of gp150, aside from a partial defect in the release of extracellular virus. Likewise, B-cell latency was established. However, the transient mononucleosis and spike in latently infected cells associated with the spread of MHV-68 to the spleen was significantly reduced in vgp150Delta-infected mice. A soluble, recombinant gp150 was found to bind specifically to B cells but not to epithelial cells in culture. In addition, gp150-deficient MHV-68 derived from mouse lungs bound less well to spleen cells than wild-type virus. Thus, gp150 is highly similar in function in vitro to the Epstein-Barr virus gp350/220. These results suggest a role for these analogous proteins in mononucleosis and have implications for their use as vaccine antigens.