国产精品国产精品一区精品国产自在现偷99精品国产在热2019国产拍偷精品网国产精品视频全国免费观看,国产精品v欧美精品v日韩精品青青精品视频国产久久国产精品久久精品国产亚洲精品国产精品国产欧美精品一区二区三区,国产精品第一页国产亚洲精品国产福利国产精品自拍国产精品视频在线观看亚洲国产精品一区二区久久国产精品国产三级国产专不,国产精品视频大陆精大陆国产国语精品2019精品国产品对白在线285年香蕉精品国产高清自在自线隔壁老王国产在线精品在线观看精品国产福利片,国产三级精品三级在专区精品国产自在现偷国产精品一区二区三区国产日韩精品欧美一区喷水亚洲精品国产精品国自产国产在线精品一区二区不卡

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 to the start of the freezing process 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 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 just sufficiently 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 a fresh tube or flask of ATCC medium 1034.
10. Incubate at 35°C with the cap screwed on tightly (incubate a test tube on a 15° horizontal slant).

Holbrook TW, et al. Activation of the alternative complement pathway by Naegleria fowleri. Infect. Immun. 30: 58-61, 1980. PubMed: 7439979

Daggett PM, Nerad TA. The biochemical identification of vahlkampfiid amoebae. J. Protozool. 30: 126-128, 1983. PubMed: 6864593

Darby CP, et al. Primary amebic meningoencephalitis. Am. J. Dis. Child. 133: 1025-1027, 1979. PubMed: 495593

Holbrook TW, Parker BW. Naegleria fowleri in chick embryos effects of embryo age and incubation temperature, and the infectivity of embryo-derived amebae for mice. Am. J. Trop. Med. Hyg. 28: 984-987, 1979. PubMed: 574367

Holbrook TW. Naegleria fowleri in the chick embryo. Trans. R. Soc. Trop. Med. Hyg. 73: 460-462, 1979. PubMed: 555075

Olomu N, et al. Demonstration of various acid hydrolases and preliminary characterization of acid phosphatase in Naegleria fowleri. J. Protozool. 33: 317-321, 1986. PubMed: 3018238