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

I;XPERIMENI.VlII
A DOT IMMUNOBINDING ASSAY FOR ETS ASSOCIATED AVI-AN
ADENOVIRUS
The objetive ofthis woft wa lo d.vclop s. c$!y for lhc qurntinc*ion oflirdl
a.tig€n d.s pr* in HISV !.@n6 ommcrcially auilrbl. in rhe nudcl
d-Ad-!!!!&! The virus w.J propr8al.d in chicken.nbryo liver (CEL) @lh. stoM in
E.gl.! mi.imum ds..tirl 6.diun (EMEM) co .i.ins 3%
':tal
oolf erun lnd anribiolis
Gtrcpiomrdn I ms/ml af,d pdicillin 1000 i'rml) Aftcr lh. lppwee of 90/. CPE, th. cells
*eE scrapcd dd pellered !t I ,000 x g lor 20 min Thc ell pellel ws sonic ed {B Boun,
Gnbtt, Cemany) for 15 s.cwitha20 kHz microprobe 75wansorpo{e.andclarificdby
@tritus.rion rt 1,000 x a lor 20 min. Th. eD.rutui ga didtua.d for 2ln !r 3?, 000 !
I
The virus pellel was rlsuspdded h 0.1% bovin snm ilbumin in Tn$bufer siline (10
mM Trjs, 150 nM Nacl, ptl ? 6) Uninr*icd cell alluE3 wcte ptocescd in ! sinild n.nn r
to. use d conkok The vn.l $pdsio. r.d dnrol dti8o3 wcr. stor.d cl ltrC lill usn
AJ!4!!!!s lalyclanal $ft rlised asain$ AAv-4 isolit. PARC-I and !.ra negativ. to
PARc-l isolcre wqe Drep.r.d (Nem er dl, 1995) The Fcsence ofantibody jn lh.t eruo
s @nnm.d nsina AOFT (Crowlq 1973).
4L@I4!4r. The infectirily orlhe virur stock wd r$.y.d in g6-well fl.r bo(od nic.olnr.
p|!l6 unns rh. CEL Th. vim tiler *.s c.lcul.t.n by th. method oi Red a|d Muerch (l9l3)
and wa qpr.$cd a TCID'
0,1JgB!t!rr!!!!4i!4r&l&!L Dd.rninotion ofs.nsitivity lryels ror AGPT lo d.r.c.
Aa and Ab ofPARcr isl{. war condud.d Ning 3rddud prolocol (Crowl., l9?3).
.t I\t inmmobiadi@ Mat lDIAl. Simihiy, d.rmin.lion of s$tivily l4els for DIA ro
deted As md Ab ofPAlC-l holaE wi6 conduclcd. Thc DIA ued w0s r modificalion oflhc
pr.vionsly d.sribed proceduos (N@m .t a/, 1995). Nitrocelldloe sh.eh ol0 45 rm Por.
sirc wc cilhc., or inio etripi of 4 o x o 5 cm dd ffik.d vilh l@d pcNil fo. th. on. lriotr
of r hc a nrig.n d o$ or pu ncncd .nd placed inro g6'wclled Dicro.irtation pht. Two-m iorolir rc