1 Protocol for Processing parasites By/ Adnan I. Al-Hindi (M.Sc.) Parasitology The Islamic University of Gaza Faculty of Science Department of Biology 28 June, 2000 Staining of parasites 28-6-2000 (Home) (H)
2 Staining and Clearing Methods for Nematodes, Cestodes and Trematodes Bring the required organism for dissection to the laboratory. After dissection, remove both stomach and intestine, put them in a saline. Trematodes and Cestodes (Regressive staining method): Specimens were left in fixative for few days (2-3 days). Then they were stained and mounted by following these steps, the worms were brought down to water through descending series of alcohol: 100 % 90 % 80 % 70 % 50 % then transferred to the haematoxylin stain until the specimen becomes purple. The staining period depends on the size of the parasite. Trematodes were small in size and required a period of 2 minutes to be stained while cestodes, which are larger, required 4 minutes or more. (Cestodes 4 minutes) (Trematodes 2 minutes) The stained parasites were then transferred to a watch glass containing tapwater and left until they become dark blue. A differentiation step followed by discarding the water and applying acid alcohol (9.9 vol. 70 % Ethyl alcohol + 0.1 vol. Conc. HcL) (Few minutes). 70 % 80 % 90 % 100 % (Ethyl alcohol)
3 The dehydrated parasites were cleared for 2 minutes in xylen and mounted in Canada balsam. Methods for Nematodes: After being fixed in hot 70 % alcohol, nematodes become fully stretched. Two or three days later, the nematodes were removed to a small watch glass containing 5 ml of the following mixture: 96 % Ethanol-------------------------------------------95 ml Glycerol--------------------------------------------------95 ml The watch glass was left half closed for one day until all alcohol had slowly evaporated, thus the parasites were in pure glycerol after 24 hours and their internal and external morphology were quite clear, nematodes were mounted on glycerin. All parasites after being examined were drawn and measured with an aid of microscope provided with a drawing tube and ocular micrometer. Photography was done using the same microscope provided with automatic photo Camera. Classification of parasites: Classification of parasites will be carried out by using the keys. End
1 Protocol for Processing parasites By/ Adnan I. Al-Hindi (M.Sc.) Parasitology The Islamic University of Gaza Faculty of Science Department of Biology 28 June, 2000 Staining of parasites 28-6-2000 (Home) (H)
2 Staining and Clearing Methods for Nematodes, Cestodes and Trematodes Bring the required organism for dissection to the laboratory. After dissection, remove both stomach and intestine, put them in a saline. Trematodes and Cestodes (Regressive staining method): Specimens were left in fixative for few days (2-3 days). Then they were stained and mounted by following these steps, the worms were brought down to water through descending series of alcohol: 100 % 90 % 80 % 70 % 50 % then transferred to the haematoxylin stain until the specimen becomes purple. The staining period depends on the size of the parasite. Trematodes were small in size and required a period of 2 minutes to be stained while cestodes, which are larger, required 4 minutes or more. (Cestodes 4 minutes) (Trematodes 2 minutes) The stained parasites were then transferred to a watch glass containing tapwater and left until they become dark blue. A differentiation step followed by discarding the water and applying acid alcohol (9.9 vol. 70 % Ethyl alcohol + 0.1 vol. Conc. HcL) (Few minutes). 70 % 80 % 90 % 100 % (Ethyl alcohol)
3 The dehydrated parasites were cleared for 2 minutes in xylen and mounted in Canada balsam. Methods for Nematodes: After being fixed in hot 70 % alcohol, nematodes become fully stretched. Two or three days later, the nematodes were removed to a small watch glass containing 5 ml of the following mixture: 96 % Ethanol-------------------------------------------95 ml Glycerol--------------------------------------------------95 ml The watch glass was left half closed for one day until all alcohol had slowly evaporated, thus the parasites were in pure glycerol after 24 hours and their internal and external morphology were quite clear, nematodes were mounted on glycerin. All parasites after being examined were drawn and measured with an aid of microscope provided with a drawing tube and ocular micrometer. Photography was done using the same microscope provided with automatic photo Camera. Classification of parasites: Classification of parasites will be carried out by using the keys. End