Biotechnology Centre of

Ho Chi Minh City.

02 June / Athena Pham

Thc Hnh Pipet Tt

Nhng li dng pipets dn n cc th nghim
khng thnh cng! 3

Mc tiu: ly nhim sang chut bng virus Cytomegalovirus LD50 ca 1.5 x

106 PFU
- Vn quan trng l ly nhim vi nng khng c qu thp, hoc qu cao
Trc khi th nghim, virus gc s phi
Lethal Dose Curve c pha long ln lt 10 ln xc nh
100 nng virus.
90 - y l qa trnh chu nh hng rt
80 nhiu ca k thut pipet
70 - Cc li dng pipet s c tch ly
Lethality %

60
10ul 10ul 10ul 10ul 10ul
50

40

30

20
100 ul 90ul 90ul 90ul 90ul 90ul
10

0
1 1.5 2
Viral Dose (x106 PFU)
Nng qu cao: loi b Th nghim vi nng va
 Nhng li dng pipets dn n cc th nghim
khng thnh cng! 4

Nhng li tch ly khi dng Pipet s dn n nng ca dung dch

gc qu thp hoc qu cao so vi mong i
iu ny dn n khng ng liu tim, lm nh hng n kt qu
th nghim
nh hng ca cc li dng pipet theo c tnh
+5% -5%
H s li H s li
Li tch ly ca ca Li tch
% pipet H s chun pipet ly %
5 105000 100000 95000 -5
10.25 11025 10000 9025 -9.75
15.76 1157.62 1000 857.37 -14.26
21.55 121.55 100 81.45 -18.55
27.63 12.76 10 7.73 -22.62
34.01 1.34 1 0.73 -26.49

Overestimated titer leads to under-inoculation

Underestimated titer leads to over-inoculation

BOTH lead to FAILURE

Evaluation and Selection 5

Hng dn thc hnh tt pipets gip ci

thin tnh chnh xc v lp li ca
nhiu ng dng khoa hc v i sng.

- nh gi v la chn
- K thut thao tc pipet
- Hiu chnh v vn hnh.
 K hoch v quy trnh lm vic 7

ng dng
hay th
nghim

Quy trnh lm
vic t mu
ban u kt d
liu cui cng

S lng mu
yu cu cho vic
thit lp d liu

nh dng cn
thit p ng
nhu cu vi
hiu qu cao

Cng c cn
thit p ng
nhu cu cng
vic

Xem xt hiu
qu/ thi gian
nh gi v la chn 8

1. nh gi tnh cht ca cht lng :

Cht lng c tnh cht tng t nh nc
Cht lng c bit: c, nht, d bay hi.

Piston vi
Dung dch
Giong m
lng
im cui
ca dung ng mao qun
dch lng Dng mt ln
Khong trng
Piston dng
u tip khng kh
mt ln
Khng c Piston seal
khong trng
khng kh
Mu Mu

Air Displacement Positive Displacement

Pipet thng thng Pipet piston
La chn 9

Air Displacement Positive Displacement ( Pos-D)

Khuyn co t Rainin 10
nh gi v la chn 11

2. nh gi phng php x l cht lng :

Pipet in t a chc nng: Pipet lp li AutoRep E Pipet lp li AutoRep S

pipet c bn, x lp li, trn, Electronic dispense Manual dispense
chun , pha long
La chn cho phng php ELISA 13

Adjustable spacer
Pipet a knh, c th thay i
Pipet a knh khong cch gia cc knh
Pipet n knh
Ti u ha cc quy trnh lm vic 15

Thi gian ch gia cc ging nh nhau

Hon thnh 96 ging ch trong vng 6s

Liquidator 96
 nh gi v la chn 17

3. nh gi lng cht lng cn x l :

Micropipet : 0.1- 2000ul Macropipet: 5,10,20 ml Pipet X Controller: x l

c lng ln mu ln
n 100ml
nh gi v la chn 18

1. nh gi tnh cht ca cht lng.

2. nh gi phng php x l cht lng.
3. nh gi lng th tch cht lng
Type of Sample solution 19

For internal use - Confidential

La chn u tip 20

La chn ng u tip tng chnh xc v lp li

Thit k
Cht lng
Ph hp

For internal use - Confidential

Thit k Fine Point 21

For internal use - Confidential

Specialty Tip Designs 22

Filter
Filter tip: u tip chng bay hi

Low-Retention: u tip chng

bm dnh

Extended Length: u tip

ni di

Wide Orifice: u tip m

rng b mt tip xc

Shaft protection (ShaftGard): u

tip bo v thn

Gel-loading (Gel-Well): u tip cho cc mu

dng gel , dng cho cc ng dng gii m AND
hay tch phn t protein
Tip Physical Quality 23

Orifice of high-quality tip

Some poor-quality tip problems

Coaxial defect Orifice with flash Tear defect

Tip Biochemical Quality 24

For internal use - Confidential

u tip ph hp 25

u tip v pipet khng kht vi nhau c

th lm gim chnh xc ca kt qu
pipet
Khuyn co: nn s dng pipet v u tip
ca cng h thng

For internal use - Confidential

K thut thao tc 28
28
K nng c bn gim thiu ri ro. 29
29

1. Ti u di th tch o
2. Ci t micometer
3. Ra u tips trc khi ht
mu
4. Gc nhng u tips
5. su khi nhng u tips
6. Thi gian nhng u tips
7. Tc ht mu
8. Nhng k thut x mu
9. Nhng nh hng ca tay
khi thao tc (lm m pipet)
S dng pipet ph hp vi th tch 30
30

Di th tch quy nh
- 10100% th tch pipets

% sai s
Th tch khuyn dng
- Thng thng 35100% th tch
- Di 35% th ph thuc vo k thut cao
10 50 100
% th tch

chnh xc ng
Th tch ti a ca
Pipette % th tch L % L () % )
L (
100% 2.0 1.5 0.030 0.7 0.014
2 L 50% 1.0 2.7 0.027 1.3 0.013
10% 0.2 12 0.024 6 0.012

Nu s dng th tch di 10% ca pipet c th nh hng n chnh xc hn 3%

Ci t Micrometer 31
31

iu chnh qu 1/3 vng so vi th tch mnh

mong mun.
Sau iu chnh ngc li v th tch cn.
Thao tc nh th vi cc th tch khc nhau,
vi cc ln khc nhau.

Thc hin thao tc ny c th gim nh hng ti 0.5%

Correctly setting the micrometer improves accuracy up to 0.5%.
Ra trc u Tips 33

Ht mu vo u tips v sau x
ngc li hoc x ra cc thi trng
ra u tp.
Ra trc u tips m bo cho tt c
cc phn pha trong c tip xc vi
dung dch. Bn cnh l trng ra Bm v nh pipton hai
hoc ba ln ra u tips
u tips
Ra trc vi mu mnh cn x

Ra trc 2 ln gip tng chnh xc ti 0.2% vi nhng mu nc.

Gc nhng u tips (u cn) 35

Gc nhng sai Gc nhng ng

Thao tc ng
Aspirating cpipette
with the th gim nh hng
perpendicular chnh
improves xc
accuracy upti 2.5%
to 0.5%,
ultramicro up to 2.5%.
 su khi nhng u tips 36

0.1 -10 L: 1-2 mm 10 -200 L: 2-3 mm 200-2000 L: 3-6 mm

Over 2000 L: 6-10 mm

Thao tc ng c th tng chnh xc ti 1%, cn vi ultra-micro ti 5%.

Thi gian ch cho pipets th tch micro 39

Vn ngm u tips trong dung dch v i t nht 1

giy sau khi ht mu ln.
Sau t t rt u tips ra khi dung dch
iu ny rt quan trng i vi mu lng ln (th
tch mu ln) v nhng mu nht cao
K thut x mu 40

1 2
C ba k thut:
1. X bn cnh ng
2. X bn trn b mt
3. X trong dung dch
Nhng im cn lu :
- Tm dng trc khi x vi
mu c nht.
- Nu x trong dung dch, cn
gi piton v tr stop th 2
cho n khi u tips c
ly ra khi dung dch. Trnh
lm mu b ht ngc vo
3 pipets.

Vic x mu ng cch c th gim nh hng ti 1%

 Tnh nht qun 42

S dng tnh thng nht (nht

qun)
- S dng pipet nhp nhng
- p lc ln piton phi ng
u
- Tc v m i

nht qun tt nht: S dng

pipet in t
- S em li tnh nht qun
ti u
- Yu cu t nht k nng
ca ngi dng
Hn ch nh hng ca vic lm m pipet khi cm 43

To avoid hand warming the pipette, use a high-quality instrument and place it on a
stand when not in use.

hn ch nh hng lm m pipet khi cm, ta s dng pipet

c cht lng cao. V dng gi treo khi khng s dng.
K thut pipet xui 44

Load tip

Touch off Hold 1s

44
Pipet ngc l g? 46

Pipet ngc l qu trnh bt u bng vic n plunger xung v tr

dng th 2, do s bao gm thm mt lng th tch d trong
qu trnh ht mu, m khng bao gm trong lc x cui.
Pipet ngc thng c s dng vo vic lin quan n mu
mu, mu ngng t v vt liu c nht.
What is Reverse Pipetting? 47
Tng quan cc li thng gp trong pipet 48

Mc nghim
trng K thut
Cch ci t Micrometer (nu vic la chn th
tch l ti u.)
Li nh
Gc nhng u tip
<1%
Ra trc u tip
Cm nm pipet (lm m pipet)
Di th tch ti u
Li trung bnh su nhng u tip.
1-5% Tc ht mu (khng ng nht)
K thut x mu
ngp u tip / gc nghing (i vi ultra-micro)
Li ln Tc ht mu (splash-up to bt kh..)
5-25% S dng pipet air displacement cho nhng mu
c bit (nht, bay hi, nng , lnh..)

Li nh l quan trng hn i vi nhng k thut hiu chun

Vi ngi dng nn hn ch nhng li trung bnh v li ln..
Tr chi nh 49

Ma st nh Lc ma st ln

im dng trong u
tip

LTS tip Traditional tip

u Tip LTS gim lc n u tip t 3kg xung cn 0.6kg

T th thao tc 50

Tc ng theo thi gian c th nh hng c, gn v dy thn kinh

For internal use - Confidential

Khuyn co t Rainin 51

u tip c thit k s dng 1 ln.

Khng nn lm sch v ti s dng v
c tnh o lng ca n s khng
cn ng tin cy.

Trnh lau hoc thm u tip bng bt k

cch no, thm ch t bn ngoi, trong
khi cht lng ang bn trong u tip.
iu ny lm cho dung dch c xu hng
chy ra khi u tip, dn n gim
chnh xc

Thng xuyn kim tra phn u pipet

xem cn kht vi u tip khng

For internal use - Confidential

Khuyn co t Rainin 52

S dng gng tay tng chnh xc

Lun treo pipet thng ng trn gi
nc / kh ngng t c th chy ra
Lun gn 1 u tip ( vi pipet 10ul hoc
nh hn) vo pipet, k c khi khng s
dng bo v phn u pipet
Vi pipet a knh: sau khi ht phi kim
tra mc cht lng trn tt c cc knh

For internal use - Confidential

Kim tra v hiu chnh 55

Module 4 & 5: Hiu chun v vn hnh

Bo tr
Kim tra nh k
Hiu chun
Cu to pipet 56

56
Bo tr pipet ti phng th nghim 57

Bo tr pipet thng xuyn tng t l t khi hiu chun ln 25- 35%

V sinh pipet
- Kim tra phn shaft, piston, seals v o-rings. V sinh
bng dung dch de-ion, lau kh ton b bn trong v bn
ngoi
- Loi b cc cht nhim bn lo ngi nh DNA v RNAse,
bng cc nhiu gii php k c dung dch 10% thuc ty
hoc RNAse Zap.
- Trnh lm sch bng cc dung mi c tnh n mn
Khuyn co:
- D khng c nt gy vn nn thay th o-ring hng nm
- Hiu chun l khng cn thit nu pipet c bo tr
thng xuyn, ngoi tr trng hp thay th piston (
bi nh cung cp)

For internal use - Confidential

Kim tra nh k 58

C 2 phng php kim tra pipet :

Gravimetric Quick Check : cn
khi lng
Leak testing: kim tra r r

Verification results specify whether calibration is necessary.

Kt qu s xc nh xem vic hiu chun c cn thit hay khng
58
T l hiu chun 59

Khuyn co:
Kim tra nh k hng ngy/ tun/ thng/ qu
ty theo nhu cu m bo chnh xc ca
th nghim
Hiu chun ti phng th nghim ca bn
m bo chnh xc v lp li ca pipet
Accuracy v Precision 63

chnh xc- Accuracy ng ( lp li)- Precision

chnh xc l lng sai khc gia gi tr l s nh gi bng s ca s thay i
trung bnh ca rt nhiu php th vi gi tr ca mt s php o lp li nh- lch
danh ngha. Mt kt qu o c gi l chun hay h s bin thin.
chnh xc khi n gn vi gi tr ng.

For internal use - Confidential

Thng s k thut ca pipet 64

chnh xc ng
Accuracy Precision
Th tch ti a % th tch L % L () % )
L (
100% 2.0 1.5 0.030 0.7 0.014
2 L 50% 1.0 2.7 0.027 1.3 0.013
10% 0.2 12 0.024 6 0.012
100% 10 1 0.1 0.4 0.04
10 L 50% 5 1.5 0.075 0.6 0.03
10% 1 2.5 0.025 1.2 0.12
100% 20 1 0.2 0.3 0.06
20 L 50% 10 1.5 0.15 0.5 0.05
10% 2 7.5 0.15 2 0.04
100% 100 0.8 0.8 0.15 0.15
100 L 50% 50 0.8 0.4 0.24 0.12
10% 10 3.5 0.35 1 0.1
100% 200 0.8 1.6 0.15 0.3
200 L 50% 100 0.8 0.8 0.25 0.25
10% 20 2.5 0.5 1 0.2
Data from manufacturers specifications: this data can vary dependent on the user technique and many other factors
Thng s k thut ca nh sn xut v ISO 8655 65

ISO 8655
Vi cc nh cung cp : l c s cho vic kim sot cht lng v cng b cht lng ca
nh cung cp
Vi cc c quan kim tra : l c s cp mt chng ch c lp
Vi ngi s dng : cho php kim tra nh k chnh xc

For internal use - Confidential

Hiu chun 66

Tiu chun nh gi : ISO

8655
Tiu chun mi trng
Nhit : [ 0,5 C], 15-
30 C
m tng i - 50%
Gi - ~ 0
T l bay hi - ~ 0
Rung - ~ 0
cao - ~ 0

For internal use - Confidential

Tiu chun cn dng hiu chun 67
 Cn v cc thit b hiu chun pipet 68

MCP2-5 XP26PC

MCP1-S

Weighing table Pipette calibration

Phn mm hiu
chun

XP Analytical & XP Micro

+ evaporation trap SAG-P +evaporation trap AX + evaporation trap