• s • A c v e w • s c f • s o g c c o • A c M o c s f • w T • s t • a • a L. tarentolae suspension c performed a tarentolae w All growth m Leishmania. completion w vitality the c exceeded, it weeks For mainta suspension c culture (TC) f faster. It is no Best results successive d occasional h growth. It is incubate TC cultivate Leis cultivation fo of the LEXSY Always contr cultures are o motile. Thes Mid-growth p of different s containing s shape) and limitations a fungal or oth Keep patient with regular They need th If you - desp sediment cel incubation in transfection Don’t centrifu by rigorous v at 2000-300 pellets easier If you cultiva aerate the cu 10 9 cells/ml. How e needs ae culture with re t 26°C. Hig ill not survive media should Hemin is l with Hemin th completed m t is possible aining LEXSY cultures in 10 flasks. Don´t ot necessary are obtaine dilution of c higher dilutio convenient t flask uprig shmania muc or transfection manuals rol appearan of drop-like s se cells are phase culture shape. Don’t such subpop remain mot nd must not her contamina t, esp. if you doubling tim heir time to g pite following ls 3 min at 2 n ventilated T uge Leishma vortexing. Th 00g is sufficie r. If required, ate LEXSY str ulture in a 1 . If you intend Jena Bioscien w to grow erobic cond egular dilutio her temperat e at 37°C be suppleme ight-sensitive he medium m edium shoul to re-add H Y strains for 0 ml LEXSY use agitated or growth-pr ed with inoc cultures of lo ns of stationa to dilute 10 ht, lowering ch longer tha n see chapte nce and mot shape, appro e most effic es always con t hesitate to pulations. Ce tile. Enhance necessarily ations may b u are used to mes of 7 h in row or to ad g these instr 2000g, resus TC flasks. Thi nia cultures he cells are s ent for sedim , prolong cen rains in bior 0 L fermenta d to use a sti nce GmbH • Loebstedte Tel.: +4 w a Leish ditions. The ons (see chap tures lower th ented with H , so Leishm must be store d be used w Hemin (and P transfection BHI mediu d cultures for romoting, to a culations dur ow cell den ary cells at e ml cultures g aeration fo an for 7 da r 4.4, for cul ility of the ce ox. 15x5 μm cient for tra ntain subpop transfect, p ells of older ed motility m be a sign o be identified b o working w n static suspe apt to new c uctions - enc spend pellet c is approach at high spee sensitive to th mentation and ntrifugation ti reactors be c ation without rrer, avoid h er Str. 80 • D-07749 J 49-3641-6285000 • F hmania cu strains can pter 3.2 of LE he growth-ra Hemin (Cat. mania must b ed in the dar within 2 wee PenStrep) and and analys m (Cat. No. r strain main add serum to ring early st nsities as th e.g. 1:100 w 1:50 on Mo or longer in ays in the sa ltivation for p ells by micro in size with ansfection pulations of n late or prese cultures get may result fr of mid grow by microscop with bacteria. ension culture onditions counter grow carefully in f was very hel ed >10.000g hese procedu d makes gen ime rather tha careful with stirring for o high sharing f Jena • Germany • e-M Fax: +49-3641-62851 ulture n be mainta EXSY manual tes and vital No ML-108) be cultivated k at 4°C. Fo eks. Howeve d to use this sis it is conv . ML-411, 41 tenance sinc o the BHI med tationary ph is may dro will not advers nday and 1 tervals betw ame medium protein expre oscopy. Cells one flagellu and platin non- or less m erve a cultur t longer and rom nutrient wth culture st py . Leishmania es and 4-5 h wth problems resh growth lpful in rescu g and don’t ures and ma ntle and quic an using hig stirring. We obtaining hig forces Mail: info@jenabioscien 0 ained as co ls). All cultiva ity significan which is ess d in the da or optimal gro er, if this she s medium fo venient to gr 12) in ventila ce cells will a dium ase. Avoid p growth. H sely affect su :20 on Frida ween passage without dilu ession see cha of mid-grow m at the flat ng on solid motile cells an re with drop d thinner (n deprivation tage. Also, a cells are pr h in agitated s with the ho medium and uing cultures resuspend c ay lyse. Cent k resuspensio her speed e found it su gh cell densi nce.com ontinuous ations are tly and L. sential for ark. After owth and elf live is r 2 more row static ted tissue age much repeated However, ubsequent ay and to es. Don’t ution. For apter 5.4 wth phase end, and d media. nd of cells p-like cells eedle-like or other bacterial, rotozoans d cultures. ost strain, d continue esp. after ell pellets rifugation on of cell fficient to ties up to