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phm cy trng, ngi ta c th s dng nhiu phng php khc nhau nh phng
php vt l, phng php gii phu bnh sinh thit v phng php ha sinh, hoc kt
hp cc phng php ny. Mi phng php c u v nhc im ring. Tuy nhin,
phng php n gin, gi thnh xt nghim r v tng i chnh xc, c ph bin
hin nay l ELISA (Enzyme Linked Immunosorbent Assay) - xt nghim hp th min
dch lin kt vi enzyme.
H Ni, 14/4/2012
Gii thiu phng php ELISA
I.
Phn loi
u nhc im:
u im:
+ n gin nht, Nhanh chng bi v ch c mt khng th v tri qua t bc
hn.
+ Phn ng ca khng th th cp c loi b.
Nhc im:
+ c hiu b gii hn v thng th khng nguyn c t nht l 2 epitope (trnh
din khng nguyn) m phng php ny ch s dng 1 khng th gn vo mt
epitope.
+ Phn ng min dch ca khng th chnh c th nh hng bt li bng cch tc
dng vi cc enzyme.
+ Phi nh du cho tng khng th chuyn bit vi tng i tng.
+ Khng c s linh hot trong la chn khng th chnh t mt th nghim khc
+ Khuch i tn hiu ti thiu.
p dng:
- Pht hin nhim HIV tr s sinh c sinh ra t m nhim HIV.
- Pht hin sm giai on mi nhim HIV (giai on ca s) hoc khi kt qu pht hin
KT khng r rng.
- Theo di din tin bnh, nh gi hiu qu iu tr thuc khng virt v trong cc mc
ch nghin cu khc.
2. Indirect ELISA (ELISA gin tip)
Phng php ny khc Direct ELISA ch khng th bt khng nguyn khng c gn
enzyme m n l mc tiu gn c hiu ca mt khng th khc (khng th ny mi l
khng th c gn vi enzyme).
Quy trnh thc hin:
u im:
+ Bc c nh khng nguyn khng c tnh c hiu nn bt k protein no cng
gn vi b mt a v vy khng th (c nng thp) phi cnh tranh vi cc
protein khc trong huyt thanh khi gn kt vi b mt ca a. Sandwich ELISA
hn ch c nhc im ny.
+ Khng th gn enzyme c th s dng nh du cho nhiu loi khng nguyn
nn tin li v kinh t hn, d dng thng mi ha.
+ Linh hot bi v cc khng th nhiu chnh c th c thc hin trong mt loi
v cng mt khng th th cp c th c s dng pht hin.
+ nhy sng c tng ln bi v mi khng th chnh cha cc epitope mt s
c th c rng buc bi cc khng th th cp , cho php khuch i tn hiu.
Nhc im:
+ c hiu ca tng khng huyt thanh l khc nhau. iu ny dn n kt qu
khc nhau gia cc th nghim v do cn phi th nghim vi nhiu khng
huyt thanh khc nhau kt qu c th tin tng c.
3. Sandwich ELISA
y l mt dng ELISA c s dng ph bin nht trong thc tin do n cho phn ng
mnh v nhy. c gi l sandwich l do kt qu th nghim c nh gi thng qua
s kt hp ca hai loi khng th l khng th bt (capture antibodies) v khng th pht
hin (detection antibodies). K thut ny cng c phn lm hai dng l Direct
sandwich ELISA (DAS-ELISA - Double antibody sandwich) v Indirect sandwich
ELISA (TAS-ELISA Triple antibody sandwich).
3.1 Sandwich ELISA trc tip
- u im: C th pht hin s khc bit nh gia cc khng nguyn nu s dng khng
th bt v khng th pht hin khc nhau.
- V phng php ny c u im hn hn nhng phng php khc m chng ti chn
phng php ny chn on bnh virus ang nghin cu.
III.
1. nh hng n nhy
2. nh hng n kt qu
Nu cc i chng m cho kt qu dng tnh th c th do s nhim t cht to
mt yu t th nghim.
Mt s thit b s dng trong phng php ELISA
IV.
-
Micropipette.
My ra khay nha.
ng dng
V.
VI.
1. V HIV/AIDS
10
11
12
khng th khng HIV, rt ngn thi gian ca s huyt thanh. Cc sinh phm dng cc
khng nguyn c cc epitop to ra cc khng th sm v c cc loi khng nguyn cho
php pht hin khng th khng virt HIV 1 & 2, cc th type virt v cc virt HIV
nhm O.
3. Ch phm HIV UNIFORM II Plus O pht hin nhim HIV
3.1.
Nguyn tc
13
4. u im v hn ch
14
u im:
+ Cho php thc hin ng thi nhiu mu. C th dng my t ng gim bt thao tc
cho ngi lm xt nghim, trnh sai st v ly nhim.
+ c kt qu bng my khng ph thuc vo ch quan ca ngi lm xt nghim.
+ C th lu cc bng kt qu v cc thng s k thut thun li cho vic kim tra nh
gi cht lng xt nghim.
+ Gi thnh xt nghim tng i r.
Hn ch:
15
Kt lun
ELISA l mt phng php c nhiu u im ni tri so vi cc phng php khc, cho
php thc hin ng thi nhiu mu, dng my t ng gim bt thao tc cho ngi lm
xt nghim, trnh sai st v ly nhim, kt qu c bng my khng ph thuc vo ch
quan ca ngi lm xt nghim nn kt qu o c c chnh xc cao. Hn na gi
thnh xt nghim r, thi gian thu c kt qu khng qu lu. Vy nn phng php
ny nn c s dng ph bin rng ri nng cao kt qu iu tr bnh, nng sut cy
trng cng nh cht lng thc phm ca x hi. c bit, ELISA gip xt nghim
HIV/AIDS cn bnh th k ca con ngi.
Trong qu trnh lm tiu lun ny, hn vn cn nhiu thiu st do hn ch v kin thc
cng nh thi gian, v vy, rt mong c gio v cc bn gip tiu lun c hon
thin hn.
Xin chn thnh cm n!
16
17
Mc Lc
Gii thiu phng php ELISA ........................................................................................................ 1
I.
1.
2.
3.
u im v hn ch. ....................................................................................................................... 2
Phn loi .......................................................................................................................................... 3
II.
1.
2.
3.
4.
III.
1.
2.
nh hng n kt qu .................................................................................................................. 9
Mt s thit b s dng trong phng php ELISA .................................................................... 9
IV.
ng dng ........................................................................................................................................... 10
V.
1.
2.
3.
Trong y hc .................................................................................................................................... 10
ng dng ca ELISA trong xt nghim HIV ............................................................................. 10
VI.
1.
V HIV/AIDS................................................................................................................................. 10
2.
Nguyn l ca phng php ELISA pht hin khng th khng HIV .................................... 11
3.
Nguyn tc ............................................................................................................................. 13
3.2.
4.
u im v hn ch ...................................................................................................................... 14
5.
Cc kt qu bt thng ................................................................................................................ 15
Kt lun ....................................................................................................................................................... 16
Ti liu tham kho ...................................................................................................................................... 17
18