最新中文字幕-精品国产99-国产精品福利在线观看-粉嫩av一区二区三区-日韩精品久久久久-美丽的姑娘国语视频-亚洲欧洲色-国产不卡毛片-三级黄毛片-免费福利在线-日韩av三区-欧美成年人网站-欧洲美女与动交ccoo-成人涩涩软件-秋霞影院午夜老牛影院-婷婷久久伊人-呦呦视频在线观看-www.天天综合-日韩精品高清视频-国产欧美综合在线-欧美日韩一区二区三区在线视频-a猛片免费播放-91九色蝌蚪国产-国产欧美日韩专区-嫩模被强到高潮呻吟不断

1. Harvest cells from a culture that is at or near peak density by centrifugation at 600 x g for 5 min. Pool the cell pellets into a single tube.
2. Adjust the concentration of cells to 2.0 x 10⁶/ml.  If the concentration is too low, centrifuge at 600 x g for 5 minutes and resuspend the cell pellet with a volume of supernatant to yield the desired concentration.
3. Prepare a 15% (v/v) sterile DMSO solution in ATCC medium 1034 as follows:  Add the required volume of DMSO to a glass screw-capped test tube and place on ice.  Allow the DMSO to solidify.  Add the required volume of refrigerated ATCC medium 1034.  Dissolve the DMSO by inverting several times.  If the DMSO solution is not prepared on ice, an exothermic reaction will occur that may precipitate certain components of the medium.
4. Mix the cell preparation and the DMSO in equal portions. Thus, the final concentration will be 10⁶ and 7.5% (v/v) DMSO. The time from the mixing of the cell preparation and DMSO stock solution before the freezing process is begun should be no less than 15 min and no longer than 60 min.
5. Dispense in 0.5 ml aliquots into 1.0 - 2.0 ml sterile plastic screw-capped cryules (special plastic vials for cryopreservation).
6. Place vials in a controlled rate freezing unit. From room temperature cool at -1°C/min to -40°C. If freezing unit can compensate for the heat of fusion, maintain rate at -1 C/min through heat of fusion. At -40°C plunge the ampules into liquid nitrogen.
7. The frozen preparations are stored in either the vapor or liquid phase of a nitrogen refrigerator.
8. To establish a culture from the frozen state place an ampule in a water bath set at 35°C. Immerse the vial enough to cover only the frozen material. Do not agitate the vial.
9. Immediately after thawing, do not leave in the water bath, aseptically remove the contents of the ampule and inoculate into 5.0 ml of fresh ATCC medium 1034.
10. Incubate the tube on a 15° horizontal at 35°C with the cap screwed on tightly.

de Jonckheere J, Voorde H. The distribution of Naegleria fowleri in man-made thermal waters. Am. J. Trop. Med. Hyg. 26: 10-15, 1977. PubMed: 842770

De Jonckheere J. Use of an axenic medium for differentiation between pathogenic and nonpathogenic Naegleria fowleri isolates. Appl. Environ. Microbiol. 33: 751-757, 1977. PubMed: 869525

De Jonckheere JF. Occurrence of Naegleria and Acanthamoeba in aquaria. Appl. Environ. Microbiol. 38: 590-593, 1979. PubMed: 539818

De Jonckheere J, van de Voorde H. Comparative study of six strains of Naegleria with special reference to nonpathogenic variants of Naegleria fowleri. J. Protozool. 24: 304-309, 1977. PubMed: 881654

De Jonckheere JF, et al. A comparative study of 14 strains of Naegleria australiensis demonstrates the existence of a highly virulent subspecies: N. australiensis italica n. spp. J. Protozool. 31: 324-331, 1984. PubMed: 6470990

De Jonckheere JF, Dierickx PJ. Determination of acid phosphatase and leucine amino peptidase activity as an identification method for pathogenic Naegleria fowleri. Trans. R. Soc. Trop. Med. Hyg. 76: 773-775, 1982. PubMed: 7164144

De Jonckheere. Naegleria australiensis sp. nov., another pathogenic Naegleria from water. Protistologica 17: 423-429, 1981.

Environ. Res. 59: 223-226, 1992.

De Jonckheere JF. Isoenzyme patterns of pathogenic and non-pathogenic Naegleria spp. using agarose isoelectric focusing. Ann. Microbiol. 133: 319-342, 1982.

De Jonckheere JF. Isoenzyme patterns of pathogenic and non-pathogenic Naegleria spp. using agarose isoelectric focusing. Ann. Microbiol. (Paris) 133: 319-342, 1982. PubMed: 6211120

van Belkum A, et al. Genotyping Naegleria spp. and Naegleria Fowleri isolates by Interepeat Polymerase Chain reaction. J. Clin. Microbiol. 30: 2595-2598, 1992. PubMed: 1400959

Van den Driessche E, et al. Primary amoebic meningoencephalitis after swimming in stream water. Lancet 2: 971-972, 1973. PubMed: 4126595

Hadas E, Kasprzak W. Comparative biochemical studies on the pathogenic and non-pathogenic amebae of the genus Naegleria. Wiad. Parazytol. 33: 25-38, 1987. PubMed: 3307159

Rivera F, et al. Pathogenic amoebae in natural thermal waters of three resorts of Hidalgo, Mexico. Environ. Res. 50: 289-295, 1989. PubMed: 2583075

Rivera F, et al. Contaminacion del liquido cefolorraquideo de un infante con sindrome de Arnold-Chiari tipo II, hidrocefalia y mielomeningocele, por Naegleria lovaniensis. Rev. Enferm. Infecc. Pediatr. 2: 91-94, 1989.

Michel R, et al. Legionella-like slender rods multiplying within a strain of Acanthamoeba sp. isolated from drinking water. Parasitol. Res. 84: 84-88, 1998. PubMed: 9491433