LOCAL

Lab Protocols

Heavy Metal Handling, Safety and Disposal (Standard Operating Protocol)
Dan Anderson (3/15/2010 tweak of 4/22/09 tweak of 12/7/07 tweak)

Read this document first. Although the heavy metal soak protocol consists simply of adding a microliter of heavy metal solution to a crystal drop, you must know the basics of heavy metal handling and disposal BEFORE you reach for your first heavy metal compound. These heavy metal rules are very similar to radioisotope rules. Whether radioisotope or heavy metals, you must be introduced to these materials. For protection of yourself and of everybody around you, you may not spontaneously jump into using them. You might as well read this document starting at item 7 because you can't properly use heavy metals without planning at the outset for their disposal.

Toxicity has been quantified (Material Safety Data Sheets) for just a few of the many heavy metal compounds in our collection. Mercurials, for example range from really toxic (mercuric cyanide) to injectable (thimerosol). Toxicity is usually not documented because nobody (or the sacrificial fish and mice) have tasted the compounds. We use these compounds hoping that they bind the target proteins, but even if they don't, they probably bind non-target proteins in humans, so they are toxic by definition. You should use the heavy metal compounds assuming that all of them are insidious toxins, and treat them with great respect.

A listing of heavy metal compounds in our collection is available at

http://www.doe-mbi.ucla.edu/local/recipes/heavy_metals

but should only be visible from inside the MBI domain. The compound numbers refer to stickers on the caps. The reagent bottles are locked up and I have the only keys. If I give you access, please search the list rather than lifting up every bottle.

Lately, most people use heavy metal stock solutions prepared by Cameron Mura or Mike Sawaya, rather than by directly handling the dry reagents as discussed in the following sections. Users of these pre-made solutions should package heavy metal contaminated tips, Kimwipes, etc. as described below (items 7-10). Cam Mura's labelling scheme followed the cap numbers in the above listing. For example, "Hg 40" means mercuric chloride. Return Cam Mura's heavy metals box to Mike Sawaya or Dan Anderson; never leave it lying around just because it's more than 4 Angstroms away from your immediate agenda.

Do not leave anything near the microscopes. Unidentified chemical-looking items left adjacent to the microscopes are assumed to be heavy metals, because that's the primary reason why 1.5 ml centrifuge tubes of chemical-looking contents arrive adjacent to this lab's microscopes. We can't dispose of unidentified chemical-looking items.

Also, if you handle heavy metals at the microscopes, please use wet Kimwipes to wipe the focus knobs and anything else your gloves might have touched (see disposal sections, below). Don't use Kimwipes on optical surfaces.

Heavy metal handling.

 

  1. Do your heavy metal work to completion and without interruption, including clean-up. Do not rush through a heavy metal experiment to get somewhere else or to answer the phone. Focus your attention on the heavy metal experiment and nothing else.

     

  2. You must handle the heavy atom compounds wearing gloves, lab coat, eye protection, long pants and closed-toe shoes. Wear gloves even to look at the containers. If you must touch doorknobs while handling heavy metals, take off the gloves, or get somebody to open the doors for you. When you take off the gloves, they go to hazardous waste, not the trash. Other than the detail of glove disposal, all this is in UCLA's Personal Protective Equipment Policy 905 (www.adminpolicies.ucla.edu/pdf/905.pdf). One part of Policy 905 says do not enter a hallway wearing gloves.

    Do heavy atom work on top of a bench protector sheet, paper side up. The sheet will catch any splatters and drips, same as in radioisotope handling. The bench protector sheet is disposed of as hazardous waste, along with the gloves, pipet tips, etc.

     

  3. If you use spatulas or glassware for heavy metals, use those that we have dedicated to heavy metals (they are locked up, too). Glassware and spatulas used for heavy metal experiments are contaminated forever. Do not return them to general circulation. Clean the heavy metal spatula with a wet Kimwipe. The contaminated Kimwipe goes into a plastic bag as dry waste (see disposal section, below). If you must use new glassware, clearly label it, and keep it completely separate from non-heavy metal glassware. If a heavy-metal contaminated item goes into general circulation, we will have to trash all the beakers and spatulas to make sure that we eliminated the dirty one. This will shut down the lab for weeks and you ought to be fired.

     

  4. Weigh heavy metal compounds on weighing paper, or into centrifuge tubes on top of weighing paper. Use the analytical balance (the one with an enclosure), not the top-loader. Many of these compounds are corrosive to the balances, in addition to poisoning all subsequent experiments and experimentalists. If you spill any, clean up immediately (or else). Heavy metal clean-up materials go to heavy metal dry waste (see disposal sections, below).

     

  5. Use only minimal amounts of heavy atom compounds. Try to keep heavy metal experiments far below 1ml, using at most a few milligrams of heavy metal compound in a stock solution (1 ml). The volume of toxic solution and disposal bulk (that we have to pay for) may be minimized by soaking the crystals on the tops of Micro Bridges or in drops hanging from cover slips, rather than in the wells of crystallization plates.

    The heavy metal container must be well labeled, using waterproof and fade-proof ink. Write something intelligible, such as "Pt in bridge C5, Hg A3". This is of direct personal interest to me because I usually get stuck cleaning up after people who have left.

     

  6. Special handling:
    Moderately volatile mercury compounds such as methyl mercuric chloride are stored in a sealed box. Open that box in the hood and handle these compounds in the hood as much as possible. That means crossing the hallway carrying an unopened but probably externally contaminated box of mercurials. This is easiest done by two people, one operating the doorknobs without gloves. Methyl mercuric chloride leaks through vacuum grease, so don't inhale adjacent to the soak container.

    We no longer have dimethylmercury or tetraethyllead (probably permanently), but we still have separate documentation for them, for the old capillary mount method:
    http://www.doe-mbi.ucla.edu/local/recipes/DMM.html
    Osmium tetroxide is also volatile. Before breaking opening an ampoule, have ready its storage vial (Pierce Chemical/Thermo "Reactivials" with teflon cap liners, the same kind of vials as used for dimethylmercury).
    Uranium and thorium compounds are radioactive, not just toxic. They are available for use only through negotiation with me (Dan Anderson), and I am going to micromanage your experiment. I have the only keys.

    Disposal of and planning for disposal of heavy metal wastes.

     

  7. The current heavy metal segregation rule says to keep mercury and lead compounds separate. Don't put lead and mercury into the same gel if you do a binding assay via native gel shift (Boggon,TJ, Shapiro,L Structure (2000)8:R143-R149). Keep track of how much of each heavy metal reagent is in each container. Each compound has to be individually listed on the disposal tag; you may not just say "mercurials." To minimize disposal costs, segregate the concentrated wastes such as stock solutions from trace contamination wastes such as Kimwipes.

     

  8. Whatever form the wastes take, one of us with the password and hazardous waste disposal training certificate have to fill in the blanks on an online "Hazardous Waste ID Tag" (otp.ucop.edu) and attach the tag to each bottle, tube or zip-closure bag. Bagged wastes have to be double-bagged. The 22x28 inch disposal bags in 219 are Fisher 12-009-15A. We have 90 days to dispose of the waste after the first atom goes in the waste container. The spirit of this regulation is that disposal has to happen while somebody living remembers what it is.

     

  9. Collect liquid heavy metal wastes into screw-cap containers to take to hazardous waste disposal. Lead and mercury should not be in the same container. Stock solutions in 1.5 ml centrifuge tubes may be placed inside larger screw-cap tubes (such as 50 ml Falcon-style tubes) for disposal.

     

  10. Heavy metals in crystallization plates.
    If the heavy metals are on cover slips on crystallization plates, place the cover slips into 50 ml conical Falcon-style centrifuge tubes (lead and mercury separate).
    If the heavy metals are in MicroBridges, pull out the bridges and place them into zip-lock plastic bags (lead and mercury separate).
    Disposal of an entire crystallization plate containing heavy metal in the wells is quick and easy but expensive because of the bulk. If a heavy metal tray has already dried out at the time of disposal, it qualifies as "dry waste", so put it into a zip-lock bag, close the bag, then double-bag.
  11.  

  12. Disposal of dry soft things: Weighing paper, paper towels, gloves, and other dry non-sharp wastes contaminated by tiny amounts of heavy metals should be placed into zip-lock bags (lead and mercury separate). Mark the bag with what's in it. This sort of waste can usually be tagged as "solid waste 99%, mercuric chloride 1%" if that describes it. If a Kimwipe has 2 grams of mercuric chloride, it has to be tagged accordingly; be accurate.
    Disposal of dry sharp things: Contaminated sharp things such as needles and cover slips should be placed in heavy metal sharps containers such as 50ml screw-cap centrifuge tubes. Do not place anything toxic in the regular sharps containers.

     

  13. Uranium and Thorium compounds are radioactive and toxic. They should be packaged for disposal as for the other metals, but their contents identified with isotope tags, and given to the Radiation Safety Office, not to the usual hazardous waste disposal people. RSO still requires segregation, so don't mix uranium and thorium.