A Riboflavin and UV Light Treatment of Platelet Products to Eliminate Bacteria

Obtain a standard apheresis human platelet product of 90 to 360 milliliters with a concentration of 0.80 to 3.8 million platelets per microliter. Allow the bag to rest for at least two hours in a platelet incubator at 22 degrees Celsius, and then gently agitate at 60 RPM until use.

Remove a 2-milliliter sample of the platelet product, and use a blood gas analyzer at 22 degrees Celsius to ensure the pH is above 6.6. Using the same sample, measure the platelet concentration with a hematological analyzer to ensure it falls in the desired range.

Next, visually inspect the platelet bag by holding it under a direct white light source. Squeeze the bag briefly, and observe the swirling pattern of the sample. If a turbid homogeneity remains before and after squeezing the bag, do not use the platelets.

With a tubing docking device, transfer the platelets to a riboflavin and UV light illumination and storage bag. Remove the empty platelet bag using a tubing sealer, and discard it as biohazardous waste.

Next, remove the light-protective outer wrap of the 500-micromolar riboflavin stock solution. Sterile-dock the riboflavin pouch onto the inlet line of the treatment and storage bag, and drain the contents into the platelet unit. Then, hang the pouch and treatment bag set from the riboflavin pouch. Gently, press residual air out of the treatment bag, and into the riboflavin bag. Rinse any residual riboflavin into the platelet pouch with a small amount of platelet product.

Allow the riboflavin and platelet solution to drain back into the treatment bag. Then, clamp the inlet line, remove the empty riboflavin bag with the tubing sealer, and discard the empty bag appropriately. Sterile-dock a 6-inch tubing line with a female syringe connector onto the inlet line of the illumination and storage bag.

Attach a 10-milliliter syringe barrel to the tubing line. Next, add 5 milliliters of the bacterial stock suspension into the syringe barrel. Then, open the inlet line clamp, and allow the bacteria to drain into the platelet product. Rinse the syringe barrel by allowing some platelet product to flow back. After rinsing, remove the syringe barrel. Attach a 30-milliliter syringe, and flush the inlet port at least twice.

After the product is well mixed, use a clean 5-milliliter syringe, and draw a 2-milliliter pre-treatment sample. Use a new syringe to remove any air introduced to the bag. Then, remove the inlet line from the treatment bag, and weigh the platelet solution.

Begin platelet treatment by first entering the operator ID, and then mount the bag on the illuminator platen. Next, enter the sample ID using a barcode reader. Enter the product type, and enter its weight into the software. Next, close the quartz clamp to automatically deliver 6.24 joules per milliliter of ultraviolet energy. Typical treatment lasts 5 to 10 minutes.

In the meantime, titer the pre-treatment sample. After UV treatment, titer a post-treatment sample to assay bacterial survival, and then titer the stock bacterial culture as a positive control. The stock culture titer should be within one log unit CFU per milliliter of its recorded titer.