Sterilizing Plastics

Depending on the sterility requirements, products are irradiated in a dose range from approx. 15 to 20 kGy, see Section 5.6.3.2. 

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Portions (100 pL) of mycelium suspension were transferred to eight sporulation agar plates with a sterile, tmncated Eppendorf tip and spread with a sterile plastic L-shaped spreader. 

The plates were incubated for 8 days at 28 °C. Then Tween-glycerol solution (7 mL) was filled onto each plate and the spores/mycehum were suspended by rigorous scraping of the agar surface again with a sterile plastic L-shaped spreader. The spore/mycehum suspension was stored in two sterile 50 mL polypropylene tubes at —80 °C until use. 

Colony forming ability of the fetal liver cells was determined in the medium comprised 1.3% methylcellulose, 4.0 mM glutamine, 10 U/ml penicillin/-streptomycin, 100 U/ml GM-CSF, 100 U/ml IL-3, 50 ng/ml stem cell factor and 10 U/ml erythropoietin in IMDM. An aliquot of 10 cells was transferred to a 35 mm sterile plastic Petri dish and incubated at 37 C in a fully humidified atmosphere of 5% CO2 in air. The final colony count was performed on day 14 of culture, the colony types being defined by general morphological criteria. 

Transfer the cells to a 25 ml sterile universal vial using a sterile plastic Pasteur pipette. 

A sterile plastic box that can accommodate six 100 ml tissue culture plates Microcentrifuge tubes Sterile pipette tips Microscope... 

Place the plates in a sterile plastic box containing some sterile tissues sprayed with sterile water containing 50 pig/ml gentamycin sulphate and 2.5 p.g/ml amphotericin B. 

Welle, F (2005), Migration of radiolysis products from radiation-sterilized plastics, Pharm. Ind., 67(8), 970-972. 

Pour off cell suspension into a sterile plastic universal container. 

Add 5 mL of solubilization buffer to each pellet and incubate at room temperature for 10-15 min to soften inclusion bodies. Try to break the pellet apart with sterile plastic pestle or by passing it through a glass homogenizer. Some insoluble material can still remain in the solution after this step. 

Gently pour off the supernatant from the centrifuged cells and resuspend the pellet in 3 mL HBSS/BSA. Using a sterile plastic pastette, carefully layer the cell suspension onto the Percoll gradient by adding to the side wall of the tube. 

The trachea and lungs are dissected free from the heart and surrounding tissue and a sterile plastic catheter is inserted into the middle portion of the trachea and secured with silk suture. 

Perform this procedure immediately prior to transfection. Add 50 xL of serum-free media such as OptiMEM I contained in a sterile plastic tube. Add 1-4 xL of TransIT-TKO reagent directly into the serum-free media. Mix thoroughly by pipetting. Add 10-50 nM siRNA (the starting recommendation is 25 nM final concentration in the well) to the diluted transfection reagent. Mix by gentle pipetting and incubate at RT for 5-20 min (see Note 12). 

Sterile 50 mL measuring cylinder, sterile plastic filter funnel. 

Once 100% cytopathic effect (CPE) is observed, freeze the flasks and their contents at -80°C and thaw them at room temperature. Transfer the content in a sterile plastic tube and centrifuge at 1500,g for 10 min to remove cell debris. Store at -80°C. 

Disposable sterile plastic transfer (Pasteur) pipets. 

Remove slides and place several in a Petri dish, or a large 100-mm, square sterile plastic dish. These do not need to be tissue culture treated, because the cells only need to grow on the slides. Allow to air-dry in a biohazard cabinet. 

For adherent cell cultures, the following protocol for HeLa cells may serve as an example. Cervical adenocarcinoma (HeLa) cells (purchased from the American Tissue and Cell Culture Corp. ATCC Manhasset, VA, USA), cell line CCL-2, were seeded in 75 cm sterile plastic cell culture flasks (Fisher Scientific) at a concentration of approximately 2 X 10 cells cm l Typically, the growth medium consisted of 20ml Dulbecco s Modified Eagle s Medium (DMEM ATCC), supplemented with 10% fetal bovine serum (FBS ATCC). To prevent bacterial contamination, 2.5ggmT of amphotericin B (ATCC) and lOOIUmT penicillin/streptomycin (ATCC) was added to the medium. The cells were incubated at 37 °C in an atmosphere containing 5% COj. 

After centrifugation of the buffy coat (blood)/ficoll mixture, carefully collect the white intermediate ring containing PBMC by using a disposable sterile plastic pipette, and transfer to a new 50 mL tube. 

Ethylene evolution. Five days after the addition of brassinosteroid to the culture media, aliquots (5 ml) of the suspension cultures were transferred to sterile plastic tubes and tightly closed. The tubes were placed on a gyrotory shaker (140 rpm) for 12 and 24 hours and kept in the dark. Then samples were withdrawn with a syringe and injected into a gas chromatograph (Hewlett Packard 5880A column lm x 3mm packed with A1203, 60/80 mesh carrier N2, combustive gas... 

CONTAINERS FOR BLOOD and BLOOD COMPONENTS Sterile plastic containers for human blood and blood components. Empty sterile containers of plasticised poly (vinyl chloride) for human blood and blood components. 

To implant a BPB, a minimally invasive procedure is foUowecL The implantation procedure can be done in a clean Procedure Room, where the patienf s implant sites can be surgically cleansed and draped with sterile towels and covered with adherent sterile plastic drapes. The implant physician scrubs his/her forearms and hands, is gowned and gloved, and wears a cap and mask. 

Aspirate the culture supernatant of the SJ9 cells and add 1 mL Insect-Xpress without supplements. Then add the Profectin /DNA mixture drop wise and incubate the cells for 24 h at 27 °C. To prevent evaporation during the incubation, the SJ9 cell-containing culture flask is placed into a sterile plastic container containing wet filter paper to increase humidity. 

Silicone grease. Ideally, sterilize the grease by placing a small amount in the bottom a glass Petri dish (never plastic it will melt) and autoclaving it. Otherwise, it can be autoclaved in an Eppendorf tube inside a glass botde and spread afterward in a sterile plastic Petri dish. 

AS 3787.1-1997 General requirements for single-use, sterile, plasticized polyvinyl chloride (PVC) packs for human blood - Single blood packs... 

Make up all stocks using sterile disposable plastic containers and pipets. Washed glass items can be contaminated with detergents that are toxic to the eggs. Filter aU concentrated stocks through 0.45-pm Milhpore filters into sterile plastic tubes. Store frozen at 20°C. 

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