It is not clear what the aims of your pollen analysis are. Are you interested in some estimate of all the species of flowers the bees are visiting just the species the bees are actively collecting pollen from. For the latter, just scopal pollen is appropriate and counts of a mnimum of 400-600 grains per bee should give you decent estimates. You don't need to destructively take whole legs or metasomal pollen loads unless you really intend to count all those thousands and thousands of grains per bee. You always will encounter some pollen species at very low levels in these counts. These could be minor pollen hosts but more likely are "contaminants" from plants visited just for nectar or even are from plants the bees haven't visited but are present on their host plants due to secondary transfer via other visitors. If you are not concerned with dietary pollen and want an indirect measure of what the bees are visiting, you have to sample pollen from wherever it occurs on the bees bodies and often deal with small amounts. Whole body washes with sonication will get just about everything but then you have to count a lot of grains to find the rare things if they are of interest On Friday, June 30, 2017> wrote: I would caution about using liquids to remove pollen grains. Bees will usually visit a wider variety of plants for nectar than for pollen, and consequently non-host pollens can often be found on the bee’s body. The best results are obtained when grains are removed directly from the scopa, greatly reducing the chance of sampling non-host pollens. Cheers, Tom From On 6/30/17 Obviously, the limb removal strategy would be partially destructive to the specimens and would limit the ability of future researchers to use these bees, but it would be beneficial for my current project. In my situation, what would you do? You don't specify what methodology you ARE using. Regardless, I would back up one step and ask the following: Have you shown, empirically, that - by not removing all the pollen grains on each specimen - you are failing to detect pollen sources you would have detected otherwise? That is, if you have no evidence that a less than perfect level of pollen removal results in such failure, then you have no demonstrable reason to engage in more difficult procedures that will impede your research. After all, if pollen ID is done not by examining every single pollen grain individually, but by taking a random subsample, then you already have a sampling protocol that will probably not be improved measurably by achieving 100% pollen removal (i.e., if there is some pollen source that is so rare that it cannot be detected by less than 100% removal, then it's very likely that you will fail to detect it ANYWAY, because its relative abundance will not go up by simply adding more total pollen grains). That being said, what I would do, if I were absolutely determined to achieve 100% removal at all costs, is to immerse each specimen in liquid in a small vial or cuvette or similar well-sealed capsule, and sonicate it to get all the pollen liberated into the liquid. The best technique for doing so might require a little trial and error (e.g., what type of container, what type of liquid). Then the problem becomes one only of how to deal with 600 or so small capsules with pollen grains in suspension. Peace, -- Doug Yanega Dept. of Entomology Entomology Research Museum Univ. of California, Riverside, CA 92521-0314 skype: dyanega phone: (951) 827-4315 (disclaimer: opinions are mine, not UCR's) http://cache.ucr.edu/~heraty/yanega.html "There are some enterprises in which a careful disorderliness is the true method" - Herman Melville, Moby Dick, Chap. 82