Indicator Vero cell line monolayers are seeded in a 4-chamber cell culture grade slide format. The slides are incubated at 36 ± 1 C in a 5% CO 2 humidified incubator until the cells show about 60% confluency. The day of procedure initiation, 0.5 ml of test article is inoculated into each of two out of the four chambers on the slide. Similarly, Mycoplasma pneumoniae and Mycoplasma arginini were used as positive controls and inoculated into two slide chambers each. Sterile culture medium served as the negative control and was inoculated into 2 slide chambers. The slide chambers were returned to the incubator for 5 days.
On the experimental completion date, the slides inoculated with the test article, positive and negative controls are taken out of the incubator, the media removed and cells fixed and stained using Hoechst stain. The slides are then covered with coverglass and mounting fluid and examined under an epi-fluorescent microscope at 600X objective lens magnifications. Mycoplasma, if present, can be visualized as fluorescent green clusters of spherical or ellipsoidal shapes around the indicator cells and in the cytoplasm around the Vero cell nuclei.
The sponsor will NOT be required to review and approve a protocol prior to study initiation. Please fill out the sample submission form below in either Word or PDF format and send along with the samples.