Storage & Handling

Frequency

• Storage temperature and shelf-life guidelines are to help in making informed decisions for maintaining the highest activity and expected shelf life.
• The rehydration instructions are for one-time rehydration of the product to the specified concentrations.
• The safe handling guidelines should be always followed when working with select agents for the personal safety of the individual working directly with the agents and all others in the surrounding areas.
• The dilution procedures are for any time dilutions are performed.

Shelf-Life and Storage Temperature for 1 mg/ml Lyophilized and Rehydrated Toxin/Toxoid

• Refer to Table 1 for the suggested guidelines for storage temperatures (˚C) and shelf life at each storage condition to retain the highest level of activity for enterotoxins and toxoids.

Rehydration of Lyophilized Enterotoxin and/or Toxoids

• Using deionized water, rehydrate to 1 mg/ml concentration.
  • NOTE: There is residual phosphate and sodium chloride in the vial to provide a physiological phosphate buffered saline solution.

Personal Safety

• Perform all enterotoxin work in an annually certified, bio-safety cabinet/hood with sash positioned at the lowest comfortable level.
• Donning Personal Protective Equipment
  • Respiratory protection* - N95 mask to avoid inhalation or ingestion.
  • Eye and face protection* – Safety goggles and/or face shields to protect eyes and face from splashes, mists, or airborne particles from contact with eyes and skin.
  • Hand protection – Chemical-resistant gloves to avoid exposure to hands and skin.
• Lab coats, body and clothing protection to avoid exposure to skin
  • Wear disposable chemical resistant lab coat that reaches below the knee.
  • Do not wear shorts, skirts, dresses that expose skin on thighs, knees or legs.
  • Do not wear open-toed shoes.
  • Wear pants and socks that cover the ankles.

*Optional when working under biological safety cabinet/hood.

• • Doffing Personal Protective Equipment
    • The doffing procedure is to be performed in sequence to avoid re-contamination.
    • Step 1: Spritz gloves with 5% hypochlorite solution** and remove N-95 mask, if worn, spritz mask with 5% hypochlorite solution.  Discard mask in plastic lined waste container.
    • Step 2: Use sani-wipes soaked with 5% hypochlorite solution to wipe down face shield and/or goggles, if used, dispose of sani-wipe.
    • Step 3: Spritz disposable lab coat and gloves with 5% hypochlorite solution.  Remove lab coat then gloves and dispose.

**Enterotoxins are proteins and can be inactivated with a 5% hypochlorite solution within 15 minutes.  Standard (off-shelf) household bleach can be made into a 1:20 solution for use.  Enterotoxins are also inactivated by autoclaving

• Waste Disposal
  • Waste Container. Wearing PPE, spritz top of container with 5% hypochlorite solution, seal well prior to autoclaving and/or removal from laboratory.  Remove from laboratory a minimum of daily.
• Enterotoxin/toxoid Inactivation
  • Autoclave conditions: 121˚C / 1 hour / liquid cycle or conditions determined during autoclave validation/thermal mapping.
• Emergency Response Equipment
  • Emergency eye wash.
  • Use emergency exit locations and protocols.

NOTE:  These are recommendations.  Always defer to individual laboratory PPE and emergency response procedures.

Dilution procedures

Diluting to a lower concentration:

• FOR EXAMPLE PURPOSES ONLY: Dilute a 1 mg/mL to a 20ng/ml concentration
• To dilute a solution from 1 mg/mL to 20 ng/mL, you need to dilute it 50,000 times. Here’s how you can calculate and perform the dilution:
• Initial concentration:
  • Initial concentration is 1 mg/mL.
• Convert the initial concentration to ng/mL:

1 mg/mL = 1,000,000 ng/mL (since 1 mg = 1,000,000 ng)

• Desired concentration:

Desired concentration is 20 ng/mL.

• Calculate the dilution factor:

Dilution factor = Initial concentration / Desired concentration

Dilution factor = 1,000,000 ng/mL / 20 ng/mL

Dilution factor = 50,000

• Perform the dilution:

• • To achieve a concentration of 20 ng/mL from 1 mg/mL, you need to dilute the initial solution by a factor of 50,000.
  • This means for every 1 unit of the initial solution, you need to add 49,999 units of the diluent (such as buffer).
  • Steps
    • Take 1 mL of the 1 mg/mL solution.
    • Add it to 49 mL of the diluent (total volume now 50 mL).
    • Mix thoroughly to get a final concentration of 20 ng/mL.