Skip to main content Skip to main navigation menu Skip to site footer
Type: Article
Published: 2020-06-12
Page range: 447–472
Abstract views: 670
PDF downloaded: 16

Stains for entomological microtechnique: simple stains for whole mounts and dissection

Institute of Science, Natural Resources & Outdoor Studies, University of Cumbria, Fusehill Street, Carlisle, UK, CA1 2HH.
General Stains preservation wholemounts photostability microscopy slides

Abstract

Slide mounted entomological specimens often require the aid of contrast techniques to improve the clarity of morphological characteristics. Methods can involve the use of techniques such as Phase contrast, Dark field or differential interference contrast microscopy (DIC), however where an entomologist may only have access to simple brightfield microscopy chemical staining of the specimen may be used to improve contrast. For whole mounts of entomological specimens, a single stain, occasionally two, are often used, in comparison to histological sections that often employ multiple stains in complex protocols. A number of authors have proposed different stains and staining methods for a number of insect groups with few considering the long term qualities of the stain, it has previously been shown that aniline dyes are prone to fading in Canada Balsam mounts, and that some stains fade even when protected from sunlight. This paper aims to summarise the knowledge of stains used for entomological specimens and provide details on the archival qualities.

 

References

  1. Barbosa, P., Berry, R. & Kary, C.K. (2014) Insect histology: Practical laboratory techniques. Wiley-Blackwell, Sussex, 368 pp.

    https://doi.org/10.1002/9781118876114

    Barr, D. (1973) Methods for the collection, preservations and study of water mites (Acari: Parasitengona). Life Sciences Miscellaneous Publications. Royal Ontario Museum, Toronto, 28 pp.

    https://doi.org/10.5962/bhl.title.60770

    Britten, H. (1930) On making preparations of small whole insects. In: Eltringham, H. Histological and illustrative methods for entomologists. Oxford University Press, Oxford, pp. 96‒104.

    Brown, P.A. (1997) A review of techniques used in the preparation, curation and conservation of microscope slides at the Natural History Museum, London. The Biology Curator, 10 (Special Supplement), 1‒33.

    Cannon, H.G. (1937) A new biological stain for general purposes. Nature, 139, 549.

    https://doi.org/10.1038/139549a0

    Cannon, H.G. (1941) On Chlorazol Black E and some other new stains. Journal of the Royal Microscopical Society, 61, 88‒94.

    https://doi.org/10.1111/j.1365-2818.1941.tb00893.x

    Chick, A.I.R. (2010) A modification of Dioni’s mountant as a substitute for Berlese mountant. Entomologist’s Monthly Magazine, 146 (2), 117‒118.

    Chick, A.I.R. (2011) A review of essential oils as clearing agents for natural history specimens. Microbehunter, 1 (11), 4‒5.

    Chick, A.I.R. (2016) Insect microscopy. Crowood Press, Wiltshire, 128 pp.

    Chick, A.I.R. (2017a) A simple method of obtaining dark-ground illumination for the examination of entomological specimens. Bulletin of the Amateur Entomology Society, 76, 20‒23.

    Chick, A.I.R. (2017b) A methodology for recovering invertebrate specimens from deteriorated gum chloral mountant media. Entomologists Monthly Magazine, 153 (2), 117‒119.

    Chick, A.I.R. (2018) A further modification of Dioni’s mounting media to allow staining, clearing and mounting of Acari. Acarologia, 58 (4), 795‒800.

    Chick, A.I.R. & Cassella, J.P. (2011) Gum damar as a substitute for Canada balsam in mounting media for microscopical specimens. Entomologists Monthly Magazine, 147 (2), 111‒114.

    Churukian, C.J. (2009) Stability of special staining solutions. Available from: https://www.urmc.rochester.edu/urmc-labs/pathology/stainsmanual/index.html?STABILITYOFSPECIALSTAININGSOLUTIONS (accessed 17 June 2019)

    Confortin, D., Neevel, H., Brustolon, M., Franco, L., Kettelarij, A.J., Williams, R.M., & van Bommel, M.R. (2010) Crystal violet: study of the photo-fading of an early synthetic dye in aqueous solution and on paper with HPLC-PDA, LC-MS and FORS. Journal of Physics: Conference Series, 2 (231), 1‒9.

    https://doi.org/10.1088/1742-6596/231/1/012011

    Colyer, C.N. & Hammond, C.O. (1951) Flies of the British Isles. Warne, London, 383 pp.

    Conn, H.J. (1953) Biological stains: A handbook on the nature and uses of the dyes employed in the biological laboratory. 6th Edition. Williams and Wilkins, Baltimore, Maryland, 380 pp.

    de Lillo, E. Craemer, C. Amrine Jr., J.W. & Nuzzaci, G. (2010) Recommended procedures and techniques for morphological studies of Eriophyoidea (Acari: Prostigmata). Experimental and Applied Acarology, 51, 283‒307.

    https://doi.org/10.1007/s10493-009-9311-x

    Dioni, W. (2006) Eosin as a nuclear stain in botany and as an economical substitute for carmine in zoology. Micscape April 2006. Available from: http://www.microscopy-uk.org.uk/mag/indexmag.html?http://www.microscopy-uk.org.uk/mag/artapr06/wd-eosin.html (accessed 17 June 2019)

    Doetschmen, W.H. (1944) Some suggestions in microtechnique particularly useful in microentomology and parasitology. Transactions of the American Microscopical Society, 63 (2), 175‒178.

    https://doi.org/10.2307/3223160

    Dublin, W.B. (1943) Combined stains method for neurologic tissue, Journal of Neuropathology & Experimental. Neurology, 2 205‒206.

    https://doi.org/10.1097/00005072-194304000-00011

    Eltringham, H. (1930) Histological and illustrative methods for entomologists. Oxford University Press, Oxford, 139 pp.

    Essig, E.O. (1948) Mounting aphids and other small insects on microscopic slides. The Pan-Pacific Entomologist, 24 (1), 9‒22.

    Evans, G.O., Sheals, J.G. & Macfarlane, D. (1961) The terrestrial Acari of the British Isles: an introduction to their morphology, biology and classification. Vol. 1. Introduction and biology. Trustees of the British Museum, London, 219 pp.

    Fairchild, G.B. & Herig, M. (1948) An improved method for mounting small insects. Science, 108, 20‒21.

    https://doi.org/10.1126/science.108.2792.20

    Faraji, F. & Bakker, F. (2008) A modified method for clearing, staining and mounting plant-inhabiting mites. European Journal of Entomology, 105, 793‒795.

    https://doi.org/10.14411/eje.2008.105

    Galigher, A.E. & Kazloff, E.N. (1971) Essentials of practical microtechnique. 2nd Edition. Lea & Febiger, Philadelphia, 531pp.

    Gates Clarke, J.F. (1941) The preparation of slides of the genitalia of Lepidoptera. Bulletin of the Brooklyn Entomological Society, 35 (4), 149‒161.

    Gray, P. (1975) The Microtomist’s formulary and guide. Robert E. Krieger, New York, 794 pp.

    Gurr, E. (1960) Encylopædia of microscopic stains. Leonard Hill Books, London, 498pp.

    Gurr, E. (1962) Staining animal tissues. Practical and theoretical. Leonard Hill Books, London, 631 pp.

    Gurr, G.T. (1952) Microscopical stains. Quekett Journal of Microscopy, 3, 419‒426.

    Halliday, R.B. (1994) Microscope slide mounting media results of informal study. Archives of Acarology List. Available from: http://www.nhm.ac.uk/hosted_sites/acarology/archive/summary.html (accessed 17 June 2019)

    Hinton, H.E. (1981) Biology of insect eggs. Vol. 1. Pergamon Press, Oxford, 473 pp.

    Hood, J.D. (1940) Microscopical whole mounts of insects. 2nd Mimeographed Edition. Cornell University, New York, 57 pp.

    Hopkin, S.P. (2007) A key to the Collembola (springtails) of Britain and Ireland. Field Studies Council Occasional Publications 111. Field Studies Council, Shrewsbury, 247 pp.

    Horobin, R.W. & Kiernan, J.A. (2002) Conn’s Biological Stains: A handbook of dyes, stains and fluorochromes for use in biology and medicine. 10th Edition. Bios Scientific Publications, Oxford, 576 pp.

    Humason, G.L. (1972) Animal tissue techniques. 3rd Edition. W.H. Freeman, San Francisco, 641 pp.

    Imms, A.D. (1929) Some methods of technique applicable to entomology. Bulletin of Entomological Research, 20, 165‒171.

    https://doi.org/10.1017/S0007485300021064

    Jennings, D.T. & Addy, N.D. (1968) A staining technique for jack-pine budworm egg masses. Journal of Economic Entomology, 61 (6), 1766.

    https://doi.org/10.1093/jee/61.6.1766

    Johnson, H. & Staub, P. (1927) A proposed new food dye. Industrial & Engineering Chemistry, 19, 497‒498.

    https://doi.org/10.1021/ie50208a027

    Kennedy, C.H. (1932) Methods for the study of the internal anatomy of insects, second mimeographed printing. Ohio State University, Columbus, Ohio, 102 pp.

    Kiefer, H.H. (1946) Isopropyl alcohol and phenol used in entomological technique. Journal of Economic Entomology, 39, 665‒666.

    https://doi.org/10.1093/jee/39.5.666

    Kirkpatrick, J. & Lendrum, A.C. (1939) A mounting medium for microscopical preparations giving good preservation of colour. Journal of Pathological Bacteriology, 49, 592–594.

    https://doi.org/10.1002/path.1700490321

    Krantz, G.W. & Walter, D.E. (2009) A manual of acarology, 3rd ed. Texas University Press. 816 pp.

    Lee, S. & Brown, R.L. (2006) A new method for preparing slide mounts of whole bodies of Microlepidoptera. Journal of Asia-Pacific Entomology, 9 (3), 249‒253.

    https://doi.org/10.1016/S1226-8615(08)60299-X

    Leston, D. (1953) Techniques: A simple method for making stained mounts. The Entomologist, 86, 254.

    Lillie, R.D. (1990) H.J. Conn’s Biological Stains. 9th Edition. Sigma Chemical Company, St. Louis, 692 pp.

    Marini, M. & Ferrari, R. (1998) A population survey of the Italian subterranean termite Reticulitermes lucifugus lucifugus Rossi in Bagnacavallo (Ravenna, Italy), using the triple mark recapture technique (TMR). Zoological Science, 15, 963‒969.

    https://doi.org/10.2108/zsj.15.963

    Martin, J.E.H. (1977) The insects and arachnids of Canada Part 1: Collecting, preparing and preserving insects, mites, and spiders. Supply and Services Canada, Quebec, 182 pp.

    McClung Jones, R. (1966) Basic Microscopic Technics. University of Chicago Press, Chicago, 334 pp.

    Neuhaus, B., Schmid, T. & Riedel, J. (2017) Collection management and study of microscope slides: Storage, profiling, deterioration, restoration procedures, and general recommendations. Zootaxa, 4322 (1), 1–173.

    https://doi.org/10.11646/zootaxa.4322.1.1

    Nagy, B.A.L. (1978) An improved rapid method that demonstrates cuticular topography, including sensilla, in arthropods. Proceedings of the Entomological Society of Ontario, 109, 75‒81.

    New, T.R. (1972) Handbooks for the identification of British insects. Vol. 1. Part 7. Psocoptera. Royal Entomological Society of London, London, 102 pp.

    Nye, W.P. (1947) A simple method of mounting aphids. The Pan-Pacific Entomologist, 23, 73‒74.

    Peterson, A. (1953) A manual of entomological techniques. 7th Edition. Edwards Brothers, Ann Arbor, Michigan, 366 pp.

    Richardson, D.T. (2001) Lignin Pink: A stain for insect integuments and wings. Balsam Post, 53, 24.

    Richardson, D.T. (2006) NBS (Northern Biological Supplies) reagents. Balsam Post, 72, 27.

    Richardson, D.T. (2014) Observations. Compiled by Gill, S. Available from: http://www.microscopy-uk.eu/diatomist/dtr.pdf (accessed 17 June 2019)

    Robinson, G.S. (1976) The preparation of slides of Lepidoptera genitalia with special reference to the microlepidoptera. The Entomologist’s Gazette, 27, 127‒132.

    Schauff, M.E. (2001) Collecting and preserving insects and mites: techniques and tools. Available from: https://www.ars.usda.gov/ARSUserFiles/80420580/CollectingandPreservingInsectsandMites/collpres.pdf (accessed 17th June 2019)

    Shelomi, M. & Kimsey, L.S. (2014) Vital staining of the stick insect digestive system identifies appendices of the midgut as novel system of excretion. Journal of Morphology, 275, 623‒633.

    https://doi.org/10.1002/jmor.20243

    Southgate, H.W. (1927) A suggested substitute for Canada Balsam as a mounting medium. Journal of the Royal Microscopical Society, 43, 311‒314.

    https://doi.org/10.1111/j.1365-2818.1923.tb05178.x

    Sukontason, K., Sukontoson, K.L., Piangjai, S., Boonchu, N,, Kurahashi, H., Hope, M. & Olson, J.K. (2004) Identification if forensically important fly eggs using a potassium permanganate staining technique. Micron, 35 (5), 391‒395.

    https://doi.org/10.1016/j.micron.2003.12.004

    Tagestad, A.D. (1976) A technique for slide preparation of aphids, scales and insect genitalia and larvae. USDA Forest Service Research Note RM, Rocky Forest and Range experiment Station, 327, 1‒3.

    Upton, M.S. & Mantle, B.L. (2010) Methods for collecting, preserving and studying insects and other terrestrial arthropods, 5th edition. The Australian Entomology Society, Miscellaneous publication No. 3. The Australian Entomology Society, Canberra, 81 pp.

    Wikipedia (2020) Merbromin. Available form: https://en.wikipedia.org/wiki/Merbromin (accessed 13 May 2020)

    Wilkey, R.F. (1962) A simplified technique for clearing, staining and permanently mounting small arthropods. Annals of the Entomological Society of America, 55 (5), 606.

    https://doi.org/10.1093/aesa/55.5.606

    Youdeowei, A. (1977) A laboratory manual of entomology. Oxford University Press, Oxford, 208 pp.