Current Trends Recommended Guidelines for Disposing of Nitrocellulose Membranes

Because of the increased use of nitrocellulose (NC) by research and clinical laboratories, concern by institutional safety officials has been expressed for the proper disposal of these membranes. Therefore, the following alternatives for disposal are recommended with the intent that the disposal method selected will conform with applicable federal, state, local, and institutional regulations and established procedures. BACKGROUND

Uses of NC Membranes. NC membranes are thin (150 um) sheets of NC polymer that have pores of highly controlled size. Typical is use of NC membranes with pore sizes of 0.45 um suitable for capturing bacteria in a filtration process.

NC also exhibits a strong affinity for both nucleic acids (DNA and RNA) and proteins (antigens and antibodies) and is, therefore, used extensively in research and clinical settings for the immobilization of nucleic acids and/or proteins for analysis. Commonly, nucleic acids or proteins are electrophoretically separated on agarose or acrylamide gels and then transferred (blotted) from the gel onto the NC. Such terms as "Southern" (transfer of DNA), "Northern" (transfer of RNA), and "Western" (transfer of protein) blots have been coined. NC sheets up to 20 x 20 cm are often used in these blotting procedures.

Additionally, large circles (up to 138 mm diameter) and sheets (up to 23 x 23 cm) are used to transfer DNA and proteins from bacteria and/or bacteriophages cultured on agar.

Properties of NC. NC membranes are manufactured by dissolving cellulose nitrate or a mixture of cellulose nitrate and cellulose acetate in a solvent and casting the resulting "honey-consistency" solution on an endless steel belt. By controlling the conditions of solvent evaporation, controlled pores are formed in the resulting polymer sheet (membrane). DISPOSAL RECOMMENDATIONS

These recommendations assume the NC has been suitably decontaminated by approved methods to eliminate any possibility of a biohazard.

Steam sterilization is the most widely employed sterilization process, and NC membranes are suitable for autoclaving. Sterilization of NC membranes is achieved at 121 C (250 F), 1 bar for 15 minutes. As an added precaution, membranes can be placed in a pan of water to eliminate any possibility of becoming dry during autoclaving. The autoclave manufacturer's instructions for use should be carefully followed.

Burial in a sanitary landfill. The easiest, safest, and least expensive method of disposing of NC membranes is burial in an approved sanitary landfill through the normal institutional waste disposal system. Radioactive membranes should be disposed of in an approved sanitary landfill established for radioactive waste. As an added precaution, the NC should be kept moist with water by sealing it in a plastic bag before disposal.

Incineration. No fire or explosion hazard will result on incineration within a well-managed, approved incineration waste-disposal system. As an added precaution, NC should be kept moist with water by sealing it in a plastic bag before final disposition in the incinerator.

Dissolution. If there is concern about disposal with general waste, the NC can be dissolved in either acid or base or in organic solvents. The resulting solution can be discarded in the normal chemical waste disposal manner as established by the institution. The waste in the container should be properly identified.

1. Dissolution in ethyl acetate. NC will completely dissolve at room temperature in ethyl acetate (CH((3))C00C((2))H((5))) within 15 minutes; 100 ml is sufficient to dissolve 600 cm((2)) of NC membrane into a pourable liquid with shaking in an enclosed container.

2. Dissolution in sodium hydroxide/ethanol. Two parts 6 molar sodium hydroxide (NaOH) should be mixed with one part 96% ethanol (C((2))H((5))OH). NC will completely dissolve within 10 minutes at room temperature; 100 ml is sufficient to dissolve 150 cm((2)), resulting in a pourable solution.

3. Dissolution in acetic acid (glacial). NC will completely dissolve in glacial acetic acid (CH((3))COOH) within 45 minutes; 100 ml is sufficient to dissolve 200 cm((2)), resulting in a pourable solution.

Reported by Schleicher & Schuell, Inc, Keene, New Hampshire; Office of Biosafety, Office of the Director, CDC.

Editorial Note

Editorial Note: Despite theoretical concerns about explosive hazards from NC membranes in autoclaves (1), practical experience demonstrates the safety of autoclaving these materials under recommended conditions of use. Precautions should be taken to ensure that autoclaves are operating properly and in accordance with manufacturers' instructions and that the NC membranes are not allowed to become dry during the autoclave cycle.

As an alternative to autoclaving, laboratorians can use an acceptable disinfection procedure, e.g., immersing the contaminated membranes in a 10% aqueous solution of household laundry bleach for 30 minutes or other chemical solutions with demonstrated microbiocidal efficacy.

Reference

1. CDC. Nitrocellulose paper used in Western blot test--fire and potential explosion hazard. MMWR 1985;34:426.

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