Mycoplasma pneumoniae Somerson et al.

1. Follow instructions as suggested for the culturing of Mollicutes:

1. Add an additional 5% heat­-inactivated fetal bovine serum to medium #988.
2. Thaw the vial and transfer its entire contents to a T-flask containing 5.0 mL of the recommended broth.  Mix well.
3. Make serial dilutions by transferring 0.5 mL from the original tube to a tube containing 5.0 mL. Repeat process by transferring 0.5 mL from the second to a third tube, etc. Dilutions are important, not only for titration purposes, but also to keep culture in varying stages of growth. Many strains will die out rapidly once acid or alkaline conditions are reached. It is recommended to prepare several dilutions from the initial tube as the cryoprotectant used in the freeze drying process often inhibits growth.
4. Use an uninoculated tube of broth to serve as a control.
5. Plates may be inoculated to check colony morphology. You can also spot each dilution on the surface of plate (4 or more/plate) to determine the number of colony-­forming units. Growth on agar may take up to 2 weeks when grown at 37°C in 5% CO₂.  

2. Mycoplasma pneumoniae strains are very slow growing and produce a very light turbidity. Growth in broth is best observed after 10 to 14 days of incubation. It usually takes at least seven days for the first T­-flasks to start showing growth. Growth is easily recognized by an indicator change from red to orange to yellow. The cells are best transferred when the medium is orange. After medium changes to yellow, cells have started to die.
3. For long term storage of Mycoplasma pneumoniae, freeze­drying or freezing is recommended. Liquid nitrogen storage is the best method. Optimally grown cells are centrifuged at 9000 rpm for 30 minutes, the supernatant poured off, and the packed cells resuspended in a smaller amount of #2611 broth. To this, add an equal amount of sterile 20% glycerol as a cryoprotectant. 

The product is provided 'AS IS' and the viability of ATCC^® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid.  Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.

This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use. Any proposed commercial use is prohibited without a license from ATCC.

While ATCC uses reasonable efforts to include accurate and up-to-date information on this product sheet, ATCC makes no warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. ATCC does not warrant that such information has been confirmed to be accurate or complete and the customer bears the sole responsibility of confirming the accuracy and completeness of any such information.

This product is sent on the condition that the customer is responsible for and assumes all risk and responsibility in connection with the receipt, handling, storage, disposal, and use of the ATCC product including without limitation taking all appropriate safety and handling precautions to minimize health or environmental risk. As a condition of receiving the material, the customer agrees that any activity undertaken with the ATCC product and any progeny or modifications will be conducted in compliance with all applicable laws, regulations, and guidelines. This product is provided 'AS IS' with no representations or warranties whatsoever except as expressly set forth herein and in no event shall ATCC, its parents, subsidiaries, directors, officers, agents, employees, assigns, successors, and affiliates be liable for indirect, special, incidental, or consequential damages of any kind in connection with or arising out of the customer's use of the product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of such materials.

Please see the material transfer agreement (MTA) for further details regarding the use of this product. The MTA is available at www.atcc.org.