. . - NIH NEWSLETTER 06 5 onud 3 nsnginu - riuuiuu 2551 VOL 5 NO 3 JULY - SEPTEMBER 2008

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CaSBols Tau% Trypan blue dye exclusion &$t.#$~-pt\ b.naBd'Launirnnrou Ihri MT4 cells ~ i lu HTLV- iOuRrVOJlaS~aRaJ (% cell reduction) , he r :h " . y l

. . ,A 4 1- carrying adult T-cell leukemia cell line ha: MOLT-4 laIgdr~ualu& ullf uRP L a a ~ d l ~ ~ r 1 5 g w o

IHTLVlllB cells 18% continuous HTLV-Ill producer (control) n?lulfi~oJR~ulW'IdilUaIM r t fl

MOLT-4 cells rraanr~arr~m8urIuXui~io~raaEi nil 2oa ~ o n a r u l ~ u + w o r R u u ~ w S ~ o Z ~ l ~ r l w r ~ t i n ~ RPMI-1640 medium [Hyclone, Utah, USA] dfj 10% ojfjJl& fefal bovine serum (FBS) [Hyclone, Utah, USA]

ran HIV % Cell reduction = A 4 xlo~ A

leisonoin MOLT-41 HTLVlllB cells b u t i u ~ r d u . , . A no. of viable cells in control , .

d i u ~ u ~ . a a i S l u ~ ~ u i r ~ u ~ ~ r a a i d ~ l o % FBS L ~ U L ~ R ? 7 B = No..oTviable cells in test well . .

iu 1% cell-free culture fluid d -80 oJfliLaa1quR rbn:: n s a ~ f fiulnrr#o HIV Iau HIV infectivity assay iinriau d.. t - , . . . ,

~ 8 % TCID 501m1 (3) nimnaou.qnmuesr~o HIV lunaean$a&'- ::.

' 8 4- J ~ a i s a ~ a i u ~ . r ~ n ' n r ~ u I w s d n a ~ u ~ . U I n ~ ~ ~ ~ ~ ~ $ u :

N~MIMT i;ludolraii wau~ur4o HIV dr i iu i r~aaia 10 rvi l ~7.g.:

~ n l ~ ~ ~ d ~ ~ d ~ ~ ~ l ~ r n ' n n ' o l (extract) ~~L&~~B:@?uGI fold dilution) ~ ~ ~ 1 ~ 1 L ~ ~ ~ l ' b ~ ~ d ~ 20% FBS ~ a o ~ a ~ : V m.rdI~ri inryubsI~rrr i Ji romuoa naobAoiu ~a R~u'LWT n'ul-~iu l ~ r J5u in~nd~Miu dn-~ iu~ ioa i~az * -' : Bao:+~oln L ~ ~ U B ~ U B I S ~ ~ ~ B ~ F ~ I O ~ R ~ I ~ ~ I ~ O J L ~ ~ L I U I U ~ I J ' nqu Zunia 96 nqu u a : l a ? 0 ~ 1 ~ ~ / ' ~ ~ - 4 - (2x10~

R15WRU polyvinylpyrrolidone 40,000 (PVP-40) l%s&~a.) uuuiiums~n"na~~"rw~~~,i;aanw~i~.'~d'1~1~b~,

U1 AZT

r r i AZT njo Zidovudine (3'-azido- n i rpnnqu $1 B'deoxythymidine ) L ~ U anti-HIV agent d ~ i U i S n ~ R ; l 37

(Inhibition of Infection) ~ ~ a : ~ u ~ a n ~ a r ~ u i ~ u a u assay (IF@. (Inhibition of replition)

g::!::',.::, '.. , , .-- . . w t h 3 arnIBr3!%Bm~7a&i%im~ n s u ~ m a ~ 1 ._ I .".I..."

. .. .. .-.-..-- WI 5 OM 3 n m p u - m u 2551

- X . . . .-. . .--- A..~..-.. _ >...... -. ~ -- . . . .

anaaain virus control (% reduction of HIV infectivity)

msr~nrr?u!wsdmuis~u'uf~'1~ir'1n' 90% auld 6a i i b.. * d urneqnaouujr.aa HIV hnaann~aoa uaznimnasuyn

mta ~$3uurfiauKun.r~iin9in 01 ~ z r

evidence for origin of thymusderived lymphocytes.

J Nail Cancer 1972; 49:891-895.

Johnson VA. "Quantitative assay for virus Infectivity."

And 'Evalution of candidate anti-HIV agents in

% Reduction of HIV infectivity = &Q xlOO C

C = TCID50lml of virus wntml

D = TClD50iml of sample

lmmunofluorescence assay (IFA) -. ?aas?%Ml HiV antigen n j u i f IFA <1b%a6 MT4

luuoiazngu il iua~uxdaR' 6 a l f l & t r k ~ d ~ w v l ~ ~ r k a + ~ i n ~ u u $ d a & m 4 T ~ d h ~ ' n UIU 10 ar-~fi ud~'1-F I%u$J HIV antibody w v e human serum d~9m1~ ~umsaralu PBS d$ 1% Bovine serum albumin bWk

'ldaudqnanc~%o uiu 1 h l u a a ~ n ~ u < i r i l a & i h ~ $30 PBS daus8u FlTC conjugated rabbit anti-Human

I ~ G drqooia 1:200 iumrazaiu PBS kii 1% Bovine

serum albumin uhril!dau d~?nrvl~iirkaa ulu 40 ulfi un:riiu16i~humsa~aiu PBS nun mounting medium

a~uua'1aC uazQ~n'?u mver slip G ~ a I ~ d ~ J ~ l n a v l i nis r~s~ua~d$an~srr.~~udv~a HIV antigen draei MT4 &u ngar Fluorescence

b" t 3irnnaouqnauuu~~4a HIV d ~au3%uiasyu la'

r-~aidsruiw 2 Zd~l16 ~lliiiivlaiu$u:~lau d ~ ~ i a ? = l J

t t ln'%nuvu ua:n~snnrraunraungu~~nisu'uo~~~i~ C U Z ~

oiuonr%aa' (active stage) rra: .r:ozw"nod!uiraa' dr tY

(latent stege) ~ s r i 7 u i ~ ~ u n i s n ' ~ n s o ~ ~ n ~ u u u ~ ~ ~ o HIV

uo~a~u'lvrsda~%az~1dnu~1w'Lun1s~%nwi~~n~a~~ riau d o : d ~ d $ n i s # n ~ o ~ 6 d ? ~ n ~ u ~ o~ms&%!Jlua~u'1w5 n?ailnmn?iul~ufib~u~nin~aad nis%~uir0uoi uar nisi%un8Bnlunu6ia'1d

vitro". In Techniques in HIV Research. Stockton

Press. U.S.A., ISBN#O-935859-76-4, page 71-

76. and page 225-237. i Ohki K, Kimura T, Ohmura K, Morikawa Y, Jones IM,

Azuma I, et al. Monoclongl antibodies to a CD4

peptide derivative which includes the region

corresponding to an immunoglobulin CDR3:

Evidence of the involvement of pre-CDR3-related

region in HIV-1 and host cell interaction

MoMular immunology. 1%2; 291391-1400.

lylifsuya H, Weinhold KJ, Furman PA, St. Clair MH,

Lehfman SN, Gallo RC, e l al. 8'-Azido-3'-

dewythymidine (BWA5OA): an ahtiviral agent that

inhibits the i n f e c t i i and Cyropalhic effect of human

T-lymphotropio virus type Illllymphadenopathy-

associated virus in vitm. Proc Natl Acad Sci LISA

1985; 82:70967100.

Nakashima H, Matsu~ T, Harada S. Kobayashi N,

Matsuda A, Ueda T et ai. Inhibition of replication

and cytopalhic effect of human T cell iymphotmpic

virus type lllllymphadenopathy-associated virus by

3'-Azido-3'-deoxythymidine in vitro. Antimicro

Agents Chemother 1986; 30:933.937.

Miyoshi I, Taguchi H, Kubonishi 1 61 al. Type C virus-

producing cell lines derived from adult T cell

leukemia. Gann Monogr 1982; 28:219-228.

Minowada J, Ohnuma T, Moore GE. Rosette fopning

human lymphoid cell lines. I. Establishment and

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