ANL-1341 IET Results Log DRAFT_OCT 3 2024.docx

INITIAL EFFICACY TEST (IET) REPORT FOR AUTOCLAVES USE FOR ON-SITE INFECTIOUS WASTE TREATMENT

Instructions for completing this document are on pages 5–9.

NAME(s) OF PERSON PERFORMING IET:

Signature Print Date

Phone number Email address Argonne Badge No.

CUSTODIAN/OWNER OF AUTOCLAVE THAT WAS TESTED:

Signature Print Date

Phone number Email address Argonne Badge No.

Autoclave(s) Location Building/Room:

Autoclave Manufacture/Model/Serial Number(s):

OPTIONAL NOTES/COMMENTS:

Table 1 - Challenge Load A: ≤ 5% w/w Low Moisture

Categories of Material Types (examples are material types; actual items may differ) Weight (grams or pounds)

1. Liquid blood/broth cultures in tubes

2. Fibers (paper towels, wipes, pads, lab coats sent off for laundering)

3. Metal (scissors, spatulas, forceps)

5. Plastics (nitrile/latex gloves, pipets, deep well, spatulas, safety glasses, plates)

6. Pathological waste (organs, biopsy/pathology specimens)

7. Glassware (agar plates, flasks, pipettes)

8. Non-woven fibers (Tyvek® gown, booties, sleeves, coat)

Total dry load weight (grams or pounds)

Total water weight (grams or pounds)

Total weight of load (grams or pounds)

Complete The Following For Tested Load A

Biological Indicator Name and Type:

Biological Indicator Lot Number: Expiration Date:

Date/time of Incubation:

Results: SCBI control not run through autoclave. (check one) |_| SCBI positive for spore growth/survival. Control test passed.

|_| SCBI negative for spore growth/survival. Control test failed.

Results: Autoclave Test Load A SCBI (check one) |_| SCBI negative for spore growth/survival. Autoclave test passed.

|_| SCBI positive for spore growth/survival. Autoclave test failed.

OPTIONAL NOTES/COMMENTS:

Table 2 - Challenge Load B: ≥ 50% w/w High Moisture

Categories of Material Types (examples are material types; actual items may differ) Weight

Liquid blood/broth cultures in tubes

Fibers (paper towels, wipes, pads, lab coats sent off for laundering)

Metal (scissors, spatulas, forceps)

Plastics (disposable pipettes, spatulas, safety glasses, agar plates)

Pathological waste (organs, biopsy/pathology specimens)

Glassware (agar plates, flasks, pipettes)

Non-woven fibers (Tyvek® gown, booties, sleeves, coat)

Total dry load weight (grams or pounds)

Total water weight (grams or pounds)

Total weight of load (grams or pounds)

Complete The Following For Tested Load B

Biological Indicator Name and Type:

Biological Indicator Lot Number: Expiration Date:

Date/time of Incubation:

Results: SCBI control not run through autoclave. (check one) |_| SCBI positive for spore growth/survival. Control test passed.

|_| SCBI negative for spore growth/survival. Control test failed.

Results: Autoclave Test Load B SCBI (check one)

OPTIONAL NOTES/COMMENTS:

Table 3 - Challenge Load B: ≥ 70% w/w Organics

Categories of Material Types (examples are material types; actual items may differ) Weight

Liquid blood/broth cultures in tubes

Fibers (paper towels, wipes, pads, lab coats sent off for laundering)

Metal (scissors, spatulas, forceps)

Plastics (disposable pipettes, spatulas, safety glasses, agar plates)

Pathological waste (organs, biopsy/pathology specimens)

Glassware (agar plates, flasks, pipettes)

Non-woven fibers (Tyvek® gown, booties, sleeves, coat)

Total dry load weight (grams or pounds)

Total water weight (grams or pounds)

Total weight of load (grams or pounds)

Complete The Following For Tested Load C

Biological Indicator Name and Type:

Biological Indicator Lot Number: Expiration Date:

Date/time of Incubation:

Results: SCBI control not run through autoclave. (check one) |_| SCBI positive for spore growth/survival. Control test passed.

|_| SCBI negative for spore growth/survival. Control test failed.

Results: Autoclave Test Load C SCBI (check one)

OPTIONAL NOTES/COMMENTS:

Contact the Biosafety Program Manager for assistance in completing the IET.

Instructions:

Step 1. Determine the autoclave cycle parameters: Select the temperature, pressure, and duration parameters suitable for treating the PIDM you will generate. A common starting point is 121C (250F), 30 psi for 60 minutes (excluding exhaust time).

Step 2. Select a spore-containing biological indicator (SCBI): Choose from the approved list in LMS-MNL-28, Biohazardous Waste Manual, Appendix C.

Step 3. Determine total autoclave capacity: Challenge loads must be equal to 100% of the autoclave's maximum manufacture’s stated capacity. Depending on the autoclave, this may take up one biohazardous waste bag, or several bags.

Step 4. Prepare challenge loads: Each of the three challenge loads (A, B, and C) must be completed, as applicable, and must include at least 5% of each of the following categories: blood/broth cultures, fibers, metals, sharps, plastics, pathological waste, glass, non-woven fibers, and bottles of liquids. Start with all dry materials so that you can add water to adjust the moisture content according to Step 7.

If your laboratory and its autoclave doesn’t treat some of the listed materials in its waste stream, mark them as “n/a” in the tables. When preparing a test load, you can use any type of object as long as it is one of the materials listed. For example, for metal use, can use an empty metal can or aluminum washers. If your PIDM doesn’t contain organic materials, challenge load C is not required.

a. Determine how many different types of materials you will mix into one bag.

b. Gather the different material types and sort by material groups.

c. Begin by estimating the total weight of a ¾ full autoclave bag containing 5% by weight of each material you will combine in the bag.

i. For example, a ¾ full 30 gallon (114 liters) autoclave bag may weigh 2,250 grams (g) (5 pounds) when full.

ii. Divide the estimate weight of the contents by 6 items (2,250 g 6 = 375 g). Add 375 g of each item to the bag. This will produce a test load with ~ 17 percent if each of the six items in the total test load.

(375 g 2,250 g) x 100 = 16.67 percent

iii. Adjust the amounts of each of the items in the test load bag until it is ¾ full confirming that at least 5 percent of the item is in the total mixture of the bag.

iv. ADD OPTION 2

d. Load as many individual bags of waste as required to reach 100 percent (%) of the manufacturer’s waste load capacity of the autoclave model being tested.

e. Mix materials content thoroughly for even distribution in the autoclave bag.

NOTE: DO NOT autoclave liquids containing hazardous chemicals (sodium hypochlorite ethanol, and so on) or radioactive materials. To do so may result in toxic and/or flammable atmospheres that can cause an illness or fire or both. Contact the Biosafety Program Manager for further guidance if you want to autoclave liquids.

Step 5. Placing the SCBI: For each of the three challenge loads (A, B, and C), place a single biological indicator with at least 1,000,000 spores of G. stearothermophilus in a sealed container, such as a glass test tube. The container’s inside diameter must be no larger than necessary to contain the SCBI. Confirm the vial contains only the SCBI. Put the container with the SCBI in the center of the bag autoclave. If more than one bag is used, place the bag with the SCBI in the center of all the bags in the autoclave.

Step 6. Determining Moisture Content of Challenge Load:

a. Load simulated biohazard waste into a biohazards bag(s) according to Step 4 to meet the challenge load items criteria listed in Table 1, 2 and 3. All items must be dry.

b. Weigh all the bags to gain the initial dry weight of all the bags combined.

c. Use Formula 1 to determine the total amount of water to add to a dry challenge load to reach the desired target of ≤ 5% (challenge load A) or ≥ 50% (challenge load B), moisture content. The final weight is the total amount of water in kilograms you must distribute between the individual bags of the applicable dry challenge load.

Formula 1:

· X: The weight of the water to be added to dry bag(s) of materials.

· MC: Desired final Moisture Content expressed as a decimal. (e.g. 5%=0.05 or 50% = 0.5)

· Dw: Dry weight of the initial dry bag of materials.

EXAMPLE: How much water must be added to a 2,250 g dry bag of materials to produce a bag with a moisture content of ≥ 50% w/w?

a. Divide both sides by 100:

b. Multiple both side by (2,250 + )

c. Distribute the 0.05 on the right side:

d. Rearrange to isolate

e. Solve for :

So add ≥ 2,250 g of water to 2,250 g dry material to obtain a MC of ≥ 50%. This means the final weight would be 4,500 grams

Step 8. Determining Percent Organics:

d. To prepare a test waste material load that is ≥ 70% organic material, you need a systematic approach to ensure that the final mixture meets the specified organic content. Here’s a step-by-step guide to achieve this:

e. Define the Components - Identify and categorize the materials available into two groups:

i. Organic materials (for example, food waste, paper, plant matter).

ii. Inorganic materials (for example, metals, glass, plastics).

f. Determine the Proportions - Calculate the amounts of organic and inorganic materials needed to achieve a mixture that is at least 70% organic by weight.

g. Use the formula - The target is for the organic content to be at least 70% of the total weight. Let:

· 𝑊total: Total weight of the mixture.

· 𝑊organic: Weight of organic materials.

· 𝑊inorganic: Weight of inorganic materials.

i. The condition is: 𝑊organic ÷ 𝑊total ≥ 0.70

· Since 𝑊total = 𝑊organic + 𝑊inorganic, the formula becomes:

ii. 𝑊organic ÷ (𝑊organic + 𝑊inorganic) ≥ 0.70

h. Solve for Worganic

i. Worganic ≥ 0.70(Worganic) + 0.70(Winorganic)

ii. Worganic - 0.70(Worganic) ≥ 0.70(Winorganic)

iii. 0.30(Worganic) ≥ 0.70(Winorganic)

iv. Worganic ≥ (0.70(Winorganic)) ÷ 0.30

v. Worganic ≥ (Winorganic)

i. Based on the derived inequality, the weight of the organic materials should be at leased times the weight of the inorganic materials.

j. Example: Assume you have 10 kg of inorganic materials. How much organic materials needs to be added to obtain ≥ 70% inorganic load?

· 𝑊organic ≥ ×10

𝑊organic ≥

k. Verify the mixture - Check the total weight and the percentage of organic material:

𝑊total = 𝑊organic + 𝑊inorganic = 23.33 kg + 10 kg = 33.33 kg

Calculate the percentage of organic material:

𝑊organic / 𝑊total×100 = (23.33 ÷ 33.33) × 100 ≈ 70%

l. Adjust as Needed

· If you must verify the organic content is strictly greater than 70%, slightly increase the amount of organic material.

· By following this method, you can accurately prepare a test waste material load that meets or exceeds the 70% organic content requirement.

Step 9. Loosely tie or tape the bags. Affix a 3M Comply™ (Thermalog™) chemical integrator strip (or equivalent). Position the test strip so it is near the center of the autoclave. Place bags into a secondary tray. Process the IET load using the autoclave cycle anticipated to be used to render all future laboratory PIDM loads non-infectious.

Step 10. After the cycle, remove the container with the biological indicator and test it according to the manufacturer’s instructions.

Step 11. Concurrently, incubate a control vial (not run in the challenge loads) with the test vial. The control vial, left unautoclaved, should show spore growth. If no growth occurs in the control vial, the test is invalid and must be repeated.

Step 12. If the biological indicators challenged in the challenge load shows growth, confirm the correctness of cycle parameters and absence of deviations. Modify the temperature, pressure, and duration if needed, and repeat the test to achieve sterile SCBIs.

Step 13. The IET is complete when all three test loads show no spore growth in the SCBI vials and all three control SCBI show spore growth. If your PIDM doesn't contain organic materials, Challenge Load C is not required.

Step 14. If operating parameters or mechanical operations change, a new IET must be performed.

Step 15. Retained a copy of the completed ANL-1341 and make available to the Illinois EPA upon request.

Associated document: LMS-MNL-28 ANL-1341(xx/xx/2024)