色噜噜精品一区二区三区,国产乱码在线精品可播放,麻豆AV无码精品一区二区,久久亚洲精精品中文字幕,久久亚洲精品国产精品,久久久久久精品免费免费999,9999国产精品欧美久久久久久,国产精品va在线播放我和闺蜜,精品久久久无码人妻中文字幕,国产亚洲美女精品久久久,99精品视频免费热播在线观看

1. Harvest cells from a culture which is at or near peak density by centrifugation at ~800 x g for 5 min.
2. Adjust concentration of cells to 2 x 10⁷/mL in fresh medium.  If the concentration is too low, centrifuge at ~800 x g for 5 minutes and resuspend the cell pellet with a volume of supernatant to yield the desired concentration.
3. While cells are centrifuging, prepare a 10% (v/v) solution of sterile DMSO in fresh medium (broth).  The DMSO solution when first prepared will warm up due to chemical heat. The solution should be allowed to return to room temperature prior to use.
4. Mix the cell preparation and the DMSO solution in equal portions. The final concentration will be 10⁷ cells/mL and 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 more than 30 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 the ampules in a Nalgene 1°C freezing apparatus.  Place the apparatus at -80°C for 1.5 to 2 hours and then plunge ampules into liquid nitrogen.  (The cooling rate in this apparatus is approximately -1°C/min.)  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. Store in either the vapor or liquid phase of a nitrogen refrigerator.
8. To thaw a frozen ampule, place it in a 35°C water bath such that the lip of the ampule remains above the water line. Thawing time is approximately 2 to 3 minutes.  Do not agitate the ampule.  Do not leave ampule in water bath after it is thawed.
9. Remove the vial from the water bath immediately after thawing.  Aseptically transfer the contents of the ampule into 10 mL of fresh ATCC medium 2736. 
10. Incubate the flask at 20-25°C with the cap screwed on tightly.
11. Maintain as described above. 

Neva FA, et al. Cutaneous leishmaniasis--a case with persistent organisms after treatment in presence of normal immune response. Am. J. Trop. Med. Hyg. 28: 467-71, 1979. PubMed: 222157

McDowell, MA, et al. Leishmania priming of human dendritic cells for CD40 ligand-induced interleukin-12p70 secretion is strain and species dependent. Infect. Immun. 70: 3994-4001, 2002. PubMed: 12117904

Joshi, PB, et al. Targeted gene deletion in Leishmania major identifies leishmanolysin (GP63) as a virulence factor. Mol. Biochem. Parasitol. 120: 33-40, 2002. PubMed: 11849703