d)       Make serial dilutions by transferring 0.5 ml from the original tube to a tube containing 4.5 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.

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2.       For long term storage of Spiroplasma sp., freeze‑drying 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 the recommended broth.  To this, add an equal amount of sterile 20% glycerol as a cryoprotectant.  This suspension is aliquoted into small plastic vials and stored at ‑70oC or below.

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.

d)       Make serial dilutions by transferring 0.5 ml from the original tube to a tube containing 4.5 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.

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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.

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To download a certificate of origin for Spiroplasma melliferum Clark et al. (33219), enter the lot number exactly as it appears on your product label or packing slip.

f)        Plates may be inoculated to check colonial morphology.  You can also spot each dilution on the surface of plate (4 or more/plate) to determine the number of colony-forming units.  However, not all strains do well on solid medium.

2.       For long term storage of Spiroplasma sp., freeze‑drying 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 the recommended broth.  To this, add an equal amount of sterile 20% glycerol as a cryoprotectant.  This suspension is aliquoted into small plastic vials and stored at ‑70oC or below.

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The certificate of origin for that lot of Spiroplasma melliferum Clark et al. (33219) is not currently available online. Complete this form to request this certificate of origin.

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b)       Using a Pasteur or 1.0 ml pipette, withdraw approximately 0.5 to 1.0 ml from a tube containing 5.0 ml.  Rehydrate the pellet.

Clark TB, et al. Spiroplasma melliferum, a new species from the honeybee (Apis mellifera). Int. J. Syst. Bacteriol. 35: 296-308, 1985.

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f)        Plates may be inoculated to check colonial morphology.  You can also spot each dilution on the surface of plate (4 or more/plate) to determine the number of colony-forming units.  However, not all strains do well on solid medium.

The certificate of analysis for that lot of Spiroplasma melliferum Clark et al. (33219) is not currently available online. Complete this form to request this certificate of analysis.

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g)       Incubate all tubes and plates under the recommended conditions and appropriate temperature.  The time necessary for growth will vary from strain to strain.  Growth on plates generally requires additional incubation.

To download a certificate of analysis for Spiroplasma melliferum Clark et al. (33219), enter the lot number exactly as it appears on your product label or packing slip.

g)       Incubate all tubes and plates under the recommended conditions and appropriate temperature.  The time necessary for growth will vary from strain to strain.  Growth on plates generally requires additional incubation.

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ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.

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If shipping to the U.S. state of Hawaii, you must provide either an import permit or documentation stating that an import permit is not required. We cannot ship this item until we receive this documentation. Contact the Hawaii Department of Agriculture (HDOA), Plant Industry Division, Plant Quarantine Branch to determine if an import permit is required.

This product sheet is not available online. We only provide this product sheet to customers who have purchased this biosafety level 3 product. If you purchased this product, please contact Technical Service for this product sheet.

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b)       Using a Pasteur or 1.0 ml pipette, withdraw approximately 0.5 to 1.0 ml from a tube containing 5.0 ml.  Rehydrate the pellet.

Witcomb RF, et al. Revised serological classification of spiroplasmas, new provisional groups and recommendations for serotyping of isolates. Curr. Microbiol. 7: 291-296, 1982.

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ATCC determines the biosafety level of a material based on our risk assessment as guided by the current edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL), U.S. Department of Health and Human Services. It is your responsibility to understand the hazards associated with the material per your organization’s policies and procedures as well as any other applicable regulations as enforced by your local or national agencies.

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ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.

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.

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