HYDROPXRICAru)II'M SYI{DROME (ANGARA DISEASE) IN ...

EXI'I;llIMTNT.NI
DEVELOTMENT OF HETEROTYPIC VACCINE A'GAINST IIPS ASSOCIATED
AVIAN ADENOWRUS
Proprg.tion orrirutB tor v..ii!. DEprnriotr
Ro(h serolypes 4 ind 8 of.vian rdonovitos (AAVJ wd. propas ed in lltinloicRuid
For propagation or AAV,HPS koldc livcr cclli rrom sp.cinc pathog.n icc (SPt)
.mbDoMld chict n cgSr {Lolmm, C.m..y) w€c n$d lor pr€paration of prinlt
cultur6lor $e popre.lion of AAv HPS isl.lc. Live. @lls wqe sDwn in polFrrrcie 150
cm2 nasks (Falcon Plasrics, Oxnard, Calil) co.r.inins 3O nl Easle's minimal escfiid mcdia
(E MEM) suDpl.m.ntcd anh lQpl. ferd olf s.tun (Flow L!bs, LJx) lnd 5 mC/t00 n
adtamicin $lfal. (SclErinS Corpor ion, Katlenh, NJ) An.. 24 hr wh.. rfie ell
nonolayer {as complct., th. m.dirm *d rcmovcd dd th...lls were inf*led vnh lhe vins
suspcnsion Followi.g vios ld$qlion aor 60 nin, $e na* *as spplied *iih E MEM,
$pplcm.ntcd wirh 2v. loc.l c.lfson, .nd i.cubd.d .. 3?qC. Wh.. cyrop hic .lfd! tu
d o! in ovq 9o"/, of th. c.ll sh*r, @ch !id! pr.pdarion w$ subjccted lo lhr.. n@e
rhaw cyclee at -20 oC, and cmlntuged at lO00 x g to sedioenl tho cell debris. Th. ccll free
supemrunt olecb virue wa lliquored lnd srored d -r0 "c
Th. d{V-HPS vinrs presot in cich c.l! lr.. susp.nsion wee scdinentcd by
centilugalion for r hr ar 60,000 x g Pell.r.d virions wer. Huspended in I mlofPBS A
disntinuous ucrose 8fldi.nt co8istane of60y. (? nD,4t % (? ml),lt% (? nl), 25 % (7
ml) rnl 15% (? ml) oa scoe in PBs M3 prcpued i. a rolydlonq ce ntus. ru& (38 5
6l) R4urp.nded litu! ws layqcd onto ii sradiot ed cenrifuged lor.l hr 100,000xg
A bdd pr6cn. .r rh. inrdfico of th. 25 dd 357. ums hy.6 w$ coll*r.d u'nng r idil.
q,tingc bdt ar 90' Thn tut.rirl6 ililly2.d for 24 hc.r 4 "C !!dGr far chmg6 ofPBS
6ins 12-14000 Mw curofdiilysis tlbing (Fih6 Scienlific. Pitlsbursh, PA) Th. didyzed
preporitionr yere enlritug.d for 2 hs ar 100,000 x a. Th. pellets werc r4u!p.nd.d in I ol