JOURNAL OF ELECTRON MICROSCOPY TECHNIQUE 15:99-100
( 1990)
Rapid Communication An Abbreviated Method for Preparing ljcodes damhi ticks for
Scanning Electron Microscopy Observation Melinda B. Rupp EM Suites, Dept. of Veterinary Biosciences, College of Veterinary Medicine, University of Illinois, Urbana, Illinois 6 1801 Past protocols specific for SEM studies involving ticks have suggested dehydrating with complex schemes requiring an initial series of ethanol in addition to a further ethanol/Freon dehydrating mixture with the final critical point drying in yet a different type of Freon (Corwin et al.. 1979; Homsher et al., 1988; Keirens et al., 1979). For investigators wishing to avoid prolonged preparatory time and costs, an efficient and simplified method is presented herein. Nymphs and larvae of unengorged Ixodes damini were initially subjected to 70% ethanol in which they were sonicated for approximately 20 minutes and subsequently dehydrated in 5 minute steps using a partial ethanol series: 75%. 95%. 100% (3X). Critical point drying (Tousimis Autosamdri-8 14) was performed utilizing Carbon dioxide as the transitional fluid. With the aid of a dissecting microscope. freshly dried samples were carefully mounted (either dorsally or ventrally) to truncated insect pins by using a 50/50 mixture of isopropanol based colloidal graphite (Electron Microscopy Sciences) and Duco cement (Devcon corporation). The head of the pin. which was opposite the sample, was secured to an aluminum mount at an angle of approximately 30 degrees with the same colloidal graphite/Duco cement adhesive. Nymphs and larvae were sputter coated twice with Au/Pd (Emscope SC500) with total deposition of metal being 20 nanometers; before each application of metal, ticks were positioned differently . Specimens were then observed under an IS1 W - 6 Scanning Electron Microscope at 9 kV and images were recorded on Kodak graphic arts film, type 4127. Intricate details of the most delicate of features, the palpi, claws, and mechanoreceptors. displayed excellent preservation of life-like qualities as a result of the condensed preparatory regime.
References Corwin. et al . (1979) An improved method for cleaning and preparing ticks for examination with the scanning electron microscope. J. Med. Entomol., 16(4): 352-353. Homsher, P.J.. Keirans, J.E., Robbins. R.G., Irwin-Pinkley, L.I.. Sonenshine. D.E. (1988) Scanning electron microscopy of ticks for systematic studies: structure of Haller's organ in eight species of the subgenus Stemalixodes of the genus lxodes (Acari: Ixodidae). J. Med. Entomol.. 25(5): 348-353. Keirens, J.E., Clifford, C.M.. Corwin, D. (1976) Zxodes sigelos , N. Sp. (Acarina: Ixodidae), A parasite of rodents in Chile, with a method for preparing ticks for examination by scanning electron microscopy. Acarologia, 18(2): 2 17-225. 0 1990 WILEY-LISS, INC.
Received January 17, 1990; accepted
In
revised form February 6, 1990
100
M.B. RUI'P
Above: 1. Spiracular plate on ventral side of nymph. x410. 2. Claw of larva. x l , 700. 3. Hypostome and Palpi of larva, ventral side. x800. 4. Sensory receptors (mechano) on leg of nymph. x750. 5. Anal groove of nymph, ventral side. 610. 6. Haller's organ and leg of larva. x920.
( 1990)
Rapid Communication An Abbreviated Method for Preparing ljcodes damhi ticks for
Scanning Electron Microscopy Observation Melinda B. Rupp EM Suites, Dept. of Veterinary Biosciences, College of Veterinary Medicine, University of Illinois, Urbana, Illinois 6 1801 Past protocols specific for SEM studies involving ticks have suggested dehydrating with complex schemes requiring an initial series of ethanol in addition to a further ethanol/Freon dehydrating mixture with the final critical point drying in yet a different type of Freon (Corwin et al.. 1979; Homsher et al., 1988; Keirens et al., 1979). For investigators wishing to avoid prolonged preparatory time and costs, an efficient and simplified method is presented herein. Nymphs and larvae of unengorged Ixodes damini were initially subjected to 70% ethanol in which they were sonicated for approximately 20 minutes and subsequently dehydrated in 5 minute steps using a partial ethanol series: 75%. 95%. 100% (3X). Critical point drying (Tousimis Autosamdri-8 14) was performed utilizing Carbon dioxide as the transitional fluid. With the aid of a dissecting microscope. freshly dried samples were carefully mounted (either dorsally or ventrally) to truncated insect pins by using a 50/50 mixture of isopropanol based colloidal graphite (Electron Microscopy Sciences) and Duco cement (Devcon corporation). The head of the pin. which was opposite the sample, was secured to an aluminum mount at an angle of approximately 30 degrees with the same colloidal graphite/Duco cement adhesive. Nymphs and larvae were sputter coated twice with Au/Pd (Emscope SC500) with total deposition of metal being 20 nanometers; before each application of metal, ticks were positioned differently . Specimens were then observed under an IS1 W - 6 Scanning Electron Microscope at 9 kV and images were recorded on Kodak graphic arts film, type 4127. Intricate details of the most delicate of features, the palpi, claws, and mechanoreceptors. displayed excellent preservation of life-like qualities as a result of the condensed preparatory regime.
References Corwin. et al . (1979) An improved method for cleaning and preparing ticks for examination with the scanning electron microscope. J. Med. Entomol., 16(4): 352-353. Homsher, P.J.. Keirans, J.E., Robbins. R.G., Irwin-Pinkley, L.I.. Sonenshine. D.E. (1988) Scanning electron microscopy of ticks for systematic studies: structure of Haller's organ in eight species of the subgenus Stemalixodes of the genus lxodes (Acari: Ixodidae). J. Med. Entomol.. 25(5): 348-353. Keirens, J.E., Clifford, C.M.. Corwin, D. (1976) Zxodes sigelos , N. Sp. (Acarina: Ixodidae), A parasite of rodents in Chile, with a method for preparing ticks for examination by scanning electron microscopy. Acarologia, 18(2): 2 17-225. 0 1990 WILEY-LISS, INC.
Received January 17, 1990; accepted
In
revised form February 6, 1990
100
M.B. RUI'P
Above: 1. Spiracular plate on ventral side of nymph. x410. 2. Claw of larva. x l , 700. 3. Hypostome and Palpi of larva, ventral side. x800. 4. Sensory receptors (mechano) on leg of nymph. x750. 5. Anal groove of nymph, ventral side. 610. 6. Haller's organ and leg of larva. x920.
