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TRANSCRIPT
Strep
Strep
Cloning Cell culture ScreeningScale-up
purifi cationStructural
& functional studies
Lead design
E. coli.
GST-tagged proteins
Schistosoma japonicumE. coli
pGEX-1 T (27-4805-01)
pGEX-6P-1 (27-4597-01)
EcoR I Sma I Sal I Xho I Not IBamH I
PreScission Protease
Leu Glu Val Leu Phe Gln Gly Pro Leu Gly Ser Pro Glu Phe Pro Gly Arg Leu Glu Arg Pro HisCTG GAA GTT CTG TTC CAG GGG CCC CTG GGA TCC CCG GAA TTC CCG GGT CGA CTC GAG CGG CCG CAT
pGEX-6P-2 (27-4598-01)
BamH I
PreScission Protease
Leu Glu Val Leu Phe Gln Gly Pro Leu Gly Ser Pro Gly Ile Pro Gly Ser Thr Arg Ala Ala Ala SerCTG GAA GTT CTG TTC CAG GGG CCC CTG GGA TCC CCA GGA ATT CCC GGG TCG ACT CGA GCG GCC GCA TCG
EcoR I Sma I Sal I Xho I Not I
pGEX-6P-3 (27-4599-01)
BamH I
PreScission Protease
Leu Glu Val Leu Phe Gln Gly Pro Leu Gly Ser Pro Asn Ser Arg Val Asp Ser Ser Gly ArgCTG GAA GTT CTG TTC CAG GGG CCC CTG GGA TCC CCG AAT TCC CGG GTC GAC TCG AGC GGC CGC
EcoR I Sma I Sal I Xho I Not I
BamH I EcoR I Sma I Sal I Xho I Not I
BamH I EcoR I Sma I Sal I Xho I Not I
BamH I EcoR I Sma I Sal I Xho I Not I
BamH I EcoR I Sma I Sal I Xho I Not I
BamH I EcoR I Sma I Sal I Xho I Not I
BamH I EcoR I Sma I Sal I Xho I Not I
EcoR I
CTG GTT CCG CGT GGA TCC CCG GAA TTC ATC GTG ACT GAC TGA CGA
BamH I
Leu Val Pro Arg Gly Ser Pro Glu Phe Ile Val Thr Asp
Thrombin Protease
Stop codons
pGEX ~4900 bp
pBR322 ori
Bal I
BspM I
Ptac
c a l
I q
Nar I
EcoR V
BssH II
BstE II Mlu I
Apa I
Tth111 I Aat II
Pst I
p4.5 AlwN I
pSj10 Bam7Stop7
pGEX-4T-2 (27-4581-01)
pGEX-5X-1 (27-4584-01)
pGEX-5X-2 (27-4585-01)
pGEX-5X-3 (27-4586-01)
pGEX-4T-1 (27-4580-01)
pGEX-4T-3 (27-4583-01)
pGEX-3X (27-4803-01)
pGEX-2TK (27-4587-01)
Leu Val Pro Arg Gly Ser Pro Gly Ile Pro Gly Ser Thr Arg Ala Ala Ala SerCTG GTT CCG CGT GGA TCC CCA GGA ATT CCC GGG TCG ACT CGA GCG GCC GCA TCG TGA
Stop codon
Ile Glu Gly Arg Gly Ile Pro Glu Phe Pro Gly Arg Leu Glu Arg Pro His Arg AspATC GAA GGT CGT GGG ATC CCC GAA TTC CCG GGT CGA CTC GAG CGG CCG CAT CGT GAC TGA
Stop codons
Ile Glu Gly Arg Gly Ile Pro Gly Ile Pro Gly Ser Thr Arg Ala Ala Ala SerATC GAA GGT CGT GGG ATC CCC GGA ATT CCC GGG TCG ACT CGA GCG GCC GCA TCG TGA
Stop codon
Ile Glu Gly Arg Gly Ile Pro Arg Asn Ser Arg Val Asp Ser Ser Gly Arg Ile Val Thr AspATC GAA GGT CGT GGG ATC CCC AGG AAT TCC CGG GTC GAC TCG AGC GGC CGC ATC GTG ACT GAC TGA
Stop codons
Leu Val Pro Arg Gly Ser Pro Glu Phe Pro Gly Arg Leu Glu Arg Pro His Arg AspCTG GTT CCG CGT GGA TCC CCG GAA TTC CCG GGT CGA CTC GAG CGG CCG CAT CGT GAC TGA
Stop codons
Leu Val Pro Arg Gly Ser Pro Asn Ser Arg Val Asp Ser Ser Gly Arg Ile Val Thr AspCTG GTT CCG CGT GGA TCC CCG AAT TCC CGG GTC GAC TCG AGC GGC CGC ATC GTG ACT GAC TGA
Stop codons
ATC GAA GGT CGT GGG ATC CCC GGG AAT TCA TCG TGA CTG ACT GACIle Glu Gly Arg Gly Ile Pro Gly Asn Ser Ser
Stop codons
Leu Val Pro Arg Gly Ser Arg Arg Ala Ser Val
Kinase
CTG GTT CCG CGT GGA TCT CGT CGT GCA TCT GTT GGA TCC CCG GGA ATT CAT CGT GAC TGAStop codons
Thrombin Protease
Thrombin Protease
Thrombin Protease
Thrombin Protease
Factor Xa Protease
Factor Xa Protease
Factor Xa Protease
Factor Xa Protease
EcoR IBamH I Sma I
EcoR IBamH I Sma I
pGEX-2T (27-4801-01)
CTG GTT CCG CGT GGA TCC CCG GGA ATT CAT CGT GAC TGA CTG ACGLeu Val Pro Arg Gly Ser Pro Gly Ile His Arg Asp
Stop codons EcoR I
Thrombin Protease
BamH I Sma I
glutathione S-transferase
Amp r
GST
-tag
ged
prot
eins
Glutathione Sepharose
* Purification modules also contain buffers
GST-tagged proteins
Products featured: GST MultiTrap FF and GST MultiTrap 4B, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
E. coli
96-well filter plates:
Sample: E. coli
Sample preparation:
Binding buffer:
Elution buffer:
Elution method:
SDS-PAGE
500
450
400
350
300
250
2000 20 40 60 80 100 120 140
Yield GST-hippocalcin, GST MultiTrap FFYield GST-hippocalcin, GST MultiTrap 4B
Incubation time (min)
Aver
age
yiel
d of
elu
ted
prot
ein
(μg)
97
66
45
30
2014
(Mr × 103)
GST-hippocalcin
LMW
mar
kers
Star
t mat
eria
lFl
owth
roug
hEl
uate
Flow
thro
ugh
Elua
teFl
owth
roug
hEl
uate
Flow
thro
ugh
Elua
teFl
owth
roug
hEl
uate
Flow
thro
ugh
Elua
te
120 min 60 min 30 min 10 min 3 min 0 min
Summary
GST
-tag
ged
prot
eins
Products featured: GST SpinTrap Purification Module, GST Detection Module
96-well plate
GST-tagged proteins
Column:
Sample: β
Binding buffer:
Elution buffer:
SDS-PAGE
DDM OG CHAPS Tween 20Triton X-100 Sarcosyl
GST-ecoKch94
67
43
(Mr × 103)
LMW
mar
kers
Flow
thro
ugh
Elua
te fr
om G
ST S
pinT
rap
Flow
thro
ugh
Elua
te fr
om G
ST S
pinT
rap
Flow
thro
ugh
Elua
te fr
om G
ST S
pinT
rap
Flow
thro
ugh
Elua
te fr
om G
ST S
pinT
rap
Flow
thro
ugh
Elua
te fr
om G
ST S
pinT
rap
Flow
thro
ugh
Elua
te fr
om G
ST S
pinT
rap
0.8
0.6
0.4
0.2
0
Solubilized materialFlowthroughEluate
Activ
ity (A
340/
min
*ml)
DDM OGCHAPS
Tween 20
Triton X-100
Sarcosyl
Summary
Acknowledgements:
GST
-tag
ged
prot
eins
Products featured: GSTrap FF, ÄKTAexplorer, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Column:
Sample: E. coli
Binding buffer:
Elution buffer:
Flow:
System:
SDS-PAGE
0
20
40
60
80
100
%Elutionbuffer
0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
ml
Wash
Elutionbuffer
2.7 mgpure GST-taggedprotein
5.0 10.0 15.0 20.0
A280
97
66
45
30
2014
(Mr × 103)
LMW
mar
kers
Star
t mat
eria
lEl
uted
frac
tion
Summary
GST-tagged proteins
Summary
Products featured: GSTrap 4B, HiLoad 16/60 Superdex 200 pg, ÄKTAxpress, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Columns:
Sample: E. coli
Sample volume:
Binding buffer (AC):
Elution buffer (AC):
Buffer (GF):
Flow rates:
System:
SDS-PAGE
500
0
1000
1500
mAU
00205 150100
AC GF
ml
A280
Large aggregates ofGST-hippocalcin
Dimers of GST-hippocalcin
150140130120110 ml
50
0
100
150
200
250
mAUA280
9766
45
30
20
14
(Mr × 103)
GST-hippocalcin
LMW
mar
kers
Star
t mat
eria
lFl
owth
roug
h (A
C)
Elut
ed fr
actio
ns o
f dim
er (G
F)
GST
-tag
ged
prot
eins
Products featured: pGEX-2T, GSTrap FF, GSTPrep FF 16/10, ÄKTAexplorer, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Columns:
Sample: E. coli
Sample volumes:
Binding buffer:
Elution buffer:
Flow rates:
System:
SDS-PAGE
Summary
mAU
1200
1000
800
600
400
200
0
A 280 Elution buffer
100
80
60
40
20
0
%B
0
Wash
Elutionbuffer
5.5 mg
10 20 30 ml
1400
1200
1000
800
600
400
200
0
A 280
100
80
60
40
20
0
%B
Wash
Elutionbuffer
20.9 mgmAU
0 50 100 150 ml
Elution bufferElution buffer
A280
mAU 1400
1200
1000
800
600
400
200
0
Elution buffer
100
80
60
40
20
0
%B
Wash
111.0 mg
0 100 200 300 400 500 600 ml
GST-DemA
9766
45
30
20
14
(Mr × 103)
LMW
mar
kers
Star
t mat
eria
l
Star
t mat
eria
l
Flow
thro
ugh
Elut
ed fr
actio
ns
Flow
thro
ugh
Elut
ed fr
actio
nsSt
art m
ater
ial
Flow
thro
ugh
Elut
ed fr
actio
ns
GSTrap FF 1 ml GSTrap FF 5 ml GSTPrep FF 16/10
GSTrap FF 1 ml GSTrap FF 5 ml GSTPrep FF 16/10
Cleavage site
Glutathione
GST
Recombinant protein
Sepharose
Cleavage enzymesFactor Xa protease:
Thrombin protease:
PreScission Protease:
GST-tagged proteins
GST
-tag
ged
prot
eins
Products featured: GSTrap FF, PreScission Protease, ÄKTAxpress
Summary
0
10
20
30
40
50
60
70
80
90
100
10 units PreScission/mg protein20 units PreScission/mg protein40 units PreScission/mg protein
2 4 6 8 10 12 14Incubation time (h)
Cle
aved
pro
tein
(%
)
0
GST-tagged proteins
Products featured: GSTrap FF, Thrombin protease, ÄKTAexplorer, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
SDS-PAGE
Summary
Column:
Sample: E. coli
Binding buffer:
Elution buffer:
Flow rate:
System:
0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
5.0 10.0 15.0 min
WashIncubation
16 hroom temp.
Wash
A280
Targetprotein
FreeGST
2.0 4.0 6.0 8.0 10.0 12.0 min
0
20
40
60
80
100
%Elutionbuffer
97
66
45
30
20
14
(Mr × 103)
LMW
mar
kers
Star
t mat
eria
l
GST-
tagg
ed p
rote
in, 1
ml c
olum
n
GST-
tagg
ed p
rote
in, 5
ml c
olum
n
GST-
free
targ
et p
rote
in, 1
6 h
clea
vage
Clea
ved
GST
tag
Thro
mbi
n pr
otea
se (2
0 U/
ml)
LMW
mar
kers
Thrombin protease cleavage,
GSTrap FF 1 ml
Target protein
GST
-tag
ged
prot
eins
Products featured: pGEX-6P, GSTrap HP, PreScission Protease, HiLoad 16/60 Superdex 75 pg, ÄKTAxpress, ExcelGel SDS gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Summary
Columns:
Sample: α
Binding/cleavage buffer (AC):
Elution buffer (AC):
Buffer (GF):
System:
SDS-PAGE
AC GF
0
500
1000
1500
2000
mAU
200 250 300
46 mg
Cleaved protein
Regeneration
A 280
ml
97
66
45
30
20
14
(Mr × 103)
LMW
mar
kers
Star
t mat
eria
lFl
owth
roug
h
Purif
ied,
clea
ved
GST-
pur
Uncl
eave
d GS
T-pu
r
pur
GST-tagged proteins
4 5 6 7 8 9 10 11 12 1 2 3
Control Anti-luciferase detection
Columns
Products featured: pGEX-6P-1, GST 96-well Detection Module
E. coli
Summary
GST
-tag
ged
prot
eins
SDS-PAGE
9766
45
30
20
14
(Mr × 103)
Soni
cate
, E. c
oli T
G1
Soni
cate
, KL4
5 ce
lls (D
naK
over
expr
essin
g)
Soni
cate
, E. c
oli +
pGE
X-5X
-1
Soni
cate
, E. c
oli +
pGE
X-5X
-Luc
exp
ress
ing
GST-
luci
fera
se
Soni
cate
, E. c
oli +
pGE
X-4T
-E7
expr
essin
g GS
T-E7
Purif
ied
GST
Pres
tain
ed M
W m
arke
r
Products featured: Anti-GST Antibody, Amersham Hybond ECL, Amersham Hyperfilm ECL, Amersham GST Western Blotting Detection Kit
E. coli E. coli
E. coli
HRP
Summary
Product Quantity Code No.
Protein expression
λ
All vectors include BL21
Purification
Detection
Tag cleavage
Related products
Product Quantity Code No.
† available by specific customer order
For more information on equipment for chromatography and/or electrophoresis, please visit www.gelifesciences.com
GST-tagged proteins
Histidine-tagged proteins
Ni Sepharose
His
tidin
e-ta
gged
pro
tein
s
Products featured: His MultiTrap HP
Summary
96-well filter plate:
Samples: E. coli
E. coli
Equilibration buffer:
Wash buffer:
Elution buffer:
Liquid handling station:
Vacuum station:
100
75
50
37
2520
15
10
1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6M M M M
Clarified bacterial extracts
Elution, row 4
Elution,row 1
Elution, row 2
Elution, row 3
Elution, row 5
Elution, row 6
Elution, row 7
(Mr × 103)
M B P B P B P B P B P B P B P B P B P B P B P B P B
B : Blank
P : Protein purification(37 kDa)
B : Blank
P : Protein purification(37 kDa)
B : Blank
P : Protein purification(37 kDa)
150
10075
50
37
2520
15
10
(Mr × 103)
Acknowledgements:
Histidine-tagged proteins
Products featured: His SpinTrap, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
97
66
45
30
20
14
(Mr × 103)
APB 7-(His)6
LMW
mar
kers
Star
t mat
eria
l, di
lute
d 1:
10
5 m
M im
idaz
ole
durin
g bi
ndin
g, d
ilute
d 1:
2
50 m
M im
idaz
ole
durin
g bi
ndin
g, d
ilute
d 1:
2
100
mM
imid
azol
e du
ring
bind
ing,
dilu
ted
1:2
200
mM
imid
azol
e du
ring
bind
ing,
dilu
ted
1:2
Eluted pool
Column:
Sample: E.coli
Binding/wash buffer:
Elution buffer:
Summary
SDS-PAGE
His
tidin
e-ta
gged
pro
tein
s
Summary
Products featured: His GraviTrap Kit, PhastSystem, PhastGel Gradient 10–15, Amersham Hybond ECL, Anti-His Antibody, ECL Mouse IgG, HRP-linked Whole Ab, Amersham High-Range Rainbow Molecular Weight Markers
E. coli
High
mol
ecul
ar w
eigh
t mar
kers
Star
t mat
eria
l, di
lute
d 1:
20
Flow
thro
ugh,
dilu
ted
1:20
Elua
te 1
Elua
te 2
Nega
tive
cont
rol (
untr
ansf
orm
ed JM
109)
220
97
66
45
20
14
30
(Mr × 103)
High
mol
ecul
ar w
eigh
t mar
kers
Star
t mat
eria
l, di
lute
d 1:
20
Flow
thro
ugh,
dilu
ted
1:20
Elua
te 1
Elua
te 2
Nega
tive
cont
rol (
untr
ansf
orm
ed JM
109)
Western blot
Column:
Sample: E.coli
Binding/wash buffer:
Elution buffer:
SDS-PAGE
Histidine-tagged proteins
Products featured: HisTrap FF crude Kit, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Summary
030252015
Wash 1 Wash 2 Wash 3Elution 1
10500
0.5
1.0
1.5
2.0
2.5
3.0
100
200
300
400
500
Imidazole concentration
(mM)
Elution volume (ml)
A280
Elution 2
Elution 3
(Mr × 103)
9766
45
30
20
14
MBP-(His)6
LMW
mar
kers
Uncl
arifi
ed sa
mpl
e, d
ilute
d 1:
10
Was
h 5
ml,
10 m
M im
idaz
ole
Was
h 1,
10
mM
imid
azol
e
Was
h 2,
30
mM
imid
azol
e
Was
h 3,
50
mM
imid
azol
e
Elut
ion
1, 1
00 m
M im
idaz
ole
Elut
ion
2, 3
00 m
M im
idaz
ole
E
lutio
n 3,
500
mM
imid
azol
e
Column:
Sample: E. coli
Flow rate:
Buffers:
Fraction size:
Equipment:
SDS-PAGE
His
tidin
e-ta
gged
pro
tein
s
Summary
Products featured: HisTrap HP, ÄKTAprime plus
Sample: E. coli
Column:
Binding buffer:
Elution buffer:
System:
5 10 15 20 25 30 35 min
2.5
2.0
1.5
1.0
0.5
0
100
80
60
40
20
0
A280Elution
Buffer %
Histidine-tagged proteins
Pichia pastoris Products featured: HisTrap FF crude, ÄKTAexplorer, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Pichia pastoris
Summary
Column:
Sample: Saccharo-myces cerevisiae P. pastoris
Binding buffer:
Elution buffer:
Flow rate:
System:
mAU
5000
4000
3000
2000
1000
0
100
50
0
0 100 200 300 400 ml
Elution Buffer %
A280
(Mr × 103)
9766
45
30
2014
(His)6-YNR064c
LMW
mar
kers
Star
t mat
eria
l
Elut
ed p
ool,
dilu
ted
1:2
Elut
ed p
ool,
dilu
ted
1:4
SDS-PAGE
His
tidin
e-ta
gged
pro
tein
s
Products featured: HisTrap HP, ÄKTAexplorer, Biacore System
E. coli.
E. coli
Summary
500
400
300
200
100
0
91.5 92 93
pool
94 95 ml92.5 93.5 94.5
Elution buffer %100
80
60
40
20
0
500
400
300
200
100
0
0 20 40 60 80 ml
Elution buffer %mAU
100
80
60
40
20
0
1
42
3 5
Pooled fraction from the purification of refolded scFv 57P antibody fragment
100
A280
A280
mAU
Sample:
Column:
Solubilizing buffer:
Denatured binding buffer:
Refolding buffer:
Native binding buffer:
Native elution buffer:
Flow rate:
System:
5000
0
10 000
15 000
20 000
-5000
Resp
onse
sig
nal (
RU)
Reference cellSample cellResponse difference after subtraction of reference cell
200 400 600 800 1000 1200 1400 1600s
SPR sensogram
Histidine-tagged proteins
Summary
Products featured: HisTrap HP, HiPrep 26/10 Desalting, RESOURCE Q, HiLoad 16/60 Superdex 75 pg, ÄKTAxpress, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
0
500
1000
1500
mAU
1100 1200 1300 1400 1500B10
11 mg
Cleaved protein
Regeneration
A280
ml
Elution buffer %
100
80
60
40
20
0
AC DS IEX GF
97
66
45
30
20
14
(Mr × 103)
Cleaved APC1040 (Mr 36 400)Uncleaved APC1040 (Mr 38 900)
LMW
mar
kers
Star
t mat
eria
lFl
owth
roug
h
Purif
ied,
clea
ved
APC1
040
afte
r AC–
DS–I
EX–G
F
Cont
rol,
uncl
eave
d AP
C104
0
Sample: E. coli
Columns: AC:
DS:
IEX:
GF:
Cleavage conditions:
AC binding buffer:
AC cleavage buffer:
AC elution buffer:
DS and IEX binding buffer:
IEX elution buffer:
GF buffer:
His
tidin
e-ta
gged
pro
tein
s
Products featured: HisTrap FF columns, HisPrep FF 16/10, ÄKTAexplorer, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Summary
Columns:
Sample: E. coli
Sample volumes:
Binding buffer:
Elution buffer:
Flow rates:
System:
SDS-PAGE
A280
0.0 50 100 150 ml
mAU4000350030002500200015001000500
0
33 mg
100
0
50
Elutionbuffer %
HisTrap FF 1 ml HisTrap FF 5 ml HisPrep FF 16/10
0.0 5.0 10.0 15.0 20.0 25.0 30.0 40.0 ml
mAU4000350030002500200015001000
5000
6.2 mg
A280
100
0
50
Elutionbuffer % 100
0
50
Elutionbuffer %
A280
mAU4000350030002500200015001000500
00 100 200 300 500400 600 ml
149 mg
700
97
66
45
30
20
14
(Mr × 103)
MBP-(His)6
LMW
mar
kers
Star
t mat
eria
l
Elut
ed p
ool H
isTra
p FF
1 m
l
Elut
ed p
ool H
isTra
p FF
5 m
l
Elut
ed p
ool H
isPre
p FF
16/
10
Histidine-tagged proteins
Product Quantity Code No.
Purification Detection
Related products
Product Quantity Code No.
For more information on Biacore systems and chromatography and/or electrophoresis systems, please visit www.gelifesciences.com
available by specific customer order
Strep-
Strep
Strep Strep
Strep
Strep
Strep-tag II proteins
Products featured: StrepTrap HP, HiLoad 16/60 Superdex 75 pg, ÄKTAprime plus, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
StrepStrep E. coli
AC step:Column: Sample: E. coli
Strep
Flow rate: Binding buffer:
Elution buffer:
GF step:Column: Sample: Flow rate: Buffer:System:
2000
1500
0
A280 mAU
1000
500
0 10 20 30 40 ml
40
50
30
60
70
0
20
10
0 10 20 30 40 50 60 ml
A280 mAU
SummaryStrep
(Histidine)6 tag Strep-tag IImCherry Full-length target protein
(Histidine)6 tag
Strep-tag II
mCherry fragment
mCherry fragment
PreScission MYG (85-87) TEV
PreScission
MYG (85-87) TEV
97
66
45
30
20
14
(Mr × 103)
SDS-PAGE
AC step GF step
Stre
p-ta
g II
prot
eins
Products featured: StrepTrap HP, ÄKTAxpress
StrepStrep Strep
Column:
Sample: Strep
Binding buffer:
Elution buffer:
Flow rates:
System:
188
98
6249
38
28
1714
60
30
(Mr × 103)
(His)6-Strep- tag II-protein
Mar
kers
Lysa
te
Flow
thro
ugh
Elut
ed ta
rget
pro
tein
poo
l
Summary
SDS-PAGE
Acknowledgements:
Strep-tag II proteins
Products featured: StrepTrap HP columns, XK 26/20 column, StrepTactin Sepharose High Performance, ÄKTAexplorer
StrepE. coli
Summary
0
1000
2000
3000
A280 mAU
0
1000
2000
3000
A587mAU
0.0 10.0 20.0 ml
= A280
= A587
= Elution buffer, %B
Columns:
Sample: Strep E. coli
Sample volumes:
Regeneration:
Binding buffer:
Elution buffer:
Flow rates:
System:
A280 mAU
A587mAU
0
1000
2000
3000
0
1000
2000
3000
0 50 100 150 ml
A280 mAU
A587mAU
0
1000
2000
3000
0
1000
2000
3000
0 200 400 600 700 ml
StrepTrap HP 1 ml StrepTrap HP 5 ml StrepTactin Sepharose High Performance XK 26/20
Stre
p-ta
g II
prot
eins
Purification
Related products
Product Quantity Code No.
For more information on chromatography and/or electrophoresis systems, please visit www.gelifesciences.com
MBP-tagged proteins
Products featured: MBPTrap HP, HiLoad 16/60 Superdex 200 pg, ÄKTAprime plus, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
δ
Summary
2000
1500
0
1000
500
0 10 20 30 40 ml
A280mAU
100
120
50
A280mAU
0
0 20 40 60 80 100 120 ml
AC step:Column: Sample: δ E. coli
Flow rate: Binding buffer:
Elution buffer:
GF step: Column: Sample: Flow rate: Buffer:
97
66
45
30
20
14
(Mr × 103)
MBP*2-paramyosin-
-SalLM
W m
arke
rsSa
mpl
e, d
ilute
d 1:
15
Flow
thro
ugh,
dilu
ted
1:15
, MBP
Trap
HP
MBP
Trap
HP
HiLo
ad S
uper
dex 1
6/60
200
pg
Eluted pool
SDS-PAGE
MBP
-tag
ged
prot
eins
Products featured: MBPTrap HP, Superdex 200 prep grade, XK 16/20, ÄKTAprime
Summary
1000
1500
2000
2500
500
A280
mAU
0
0 20 40 60 80 100 ml120
200
300
400
100
0
0 20 40 60 80 ml
A280
mAU
170130
9572
56
43
34
26
1117
(Mr × 103)
MBP-MCAD
Mol
ecul
ar w
eigh
t mar
kers
Star
t mat
eria
l, di
lute
d 1:
6
Flow
thro
ugh
MBP
Trap
HP,
dilu
ted
1:6
Elut
ed fr
actio
ns fr
om M
BPTr
ap H
PEl
uted
frac
tions
from
Sup
erde
x 200
pg
AC stepColumn: Sample: E. coli
Binding buffer:
Elution buffer: Flow rate: System:
GF stepColumn: Sample: Buffer:Flow rate: System:
SDS-PAGE
Acknowledgements:
MBP-tagged proteins
Products featured: MBPTrap HP, HiLoad 16/60 Superdex 200 pg, ÄKTAxpress
Summary
AC column:
Sample: E. coli
Flow rate:
Binding buffer:
Elution buffer:
GF column:
Sample:
Flow rate:
Buffer:
System:
Elution buffer%B
0
500
1000
1500
2000
A280mAU
0
20
40
60
80
100
60 80 100 120 140 160 180 mlAC GF
SDS-PAGE
Acknowledgements:
250130100
70
55
35
27
15
10
(Mr × 103)
Mol
ecul
ar
wei
ght m
arke
r Eluted fractions from gel filtration
MBP
-tag
ged
prot
eins
Products featured: MBPTrap HP columns, XK 26/20 column, Dextrin Sepharose High Performance, ÄKTAexplorer, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
β
Summary
Columns:
Sample: β E. coli
Sample volumes:
Binding buffer:
Elution buffer:
Flow rates:
System:
0
500
1000
1500
2000
2500
3000
A280mAU
0.0 5.0 10.0 15.0 20.0 ml
80
100
Elution Buffer %
60
40
20
0 0
500
1000
1500
2000
2500
3000
0 100 200 300 400 500 ml
80
100
60
40
20
0600
A280mAU
Elution Buffer %
97
66
45
30
20
14
(Mr × 103)
MBP2*- -galactosidase
LMW
mar
kers
MBP
Trap
HP
1 m
l, di
lute
d 1:
6
MBP
Trap
HP
5 m
l, di
lute
d 1:
12
XK 2
6/20
, dilu
ted
1:12
MBP
Trap
HP
1 m
l, di
lute
d 1:
3
MBP
Trap
HP
5 m
l, di
lute
d 1:
3
XK 2
6/20
, dilu
ted
1:3
Star
t mat
eria
l, di
lute
d 1:
3
FlowthroughEluted pool
MBPTrap HP 1 ml Dextrin Sepharose High Performance XK 26/20
SDS-PAGE
MBP-tagged proteins
Purification
Purification
Product Quantity Code No.
For more information on Biacore systems and chromatography and/or electrophoresis systems, please visit www.gelifesciences.com
Dual-tagged proteinsProducts featured: HisTrap HP, GSTrap 4B, ÄKTAxpress, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Summary
Sample: E.coli
Columns:
Affinity chromatography:
Desalting:
Buffers:
HisTrap HP
Binding buffer:
Elution buffer:
GSTrap 4B:
Binding buffer:
Elution buffer:
HiPrep 26/10 Desalting:
Buffer:
HiPrep™
16/10
HiPrep™
16/10
HiPrep™
16/10
HiPrep™
16/10
Run 1 Run 2 Run 3
HisTrap HP HiPrep 26/10 Desalting
GSTrap 4B HiPrep 26/10 Desalting
GSTrap 4B HiPrep 26/10 Desalting
HisTrap HP HiPrep 26/10 Desalting
96
66
45
30
20
14
(Mr × 103)
LMW
mar
kers
Flow
thro
ugh
run
1El
utio
n ru
n 1
Flow
thro
ugh
run
2El
utio
n ru
n 2
Flow
thro
ugh
run
3El
utio
n ru
n 3
(His)6-GFP-GST
SDS-PAGE
Dua
l-tag
ged
prot
eins
Products featured: HisTrap HP, StrepTrap HP, ÄKTAxpress
Strep E. coli
Summary
188
9862
49
38
28
(Mr × 103)
1714
6
3
Dual-tagged Strep-tag II-(His)6
Mol
ecul
ar w
eigh
t mar
ker
Flow
thro
ugh
Elut
ed p
ool
Flow
thro
ugh
Elut
ed p
ool
Elut
ed p
ool
Flo
wth
roug
h
Individual HisTrap HP
HisTrap HP + StrepTrap HP
Individual StrepTrap HP
Sample: Strep
E. coli
Individual HisTrap HP purification
Column:
Binding buffer:
Elution buffer:
Flow rate:
Individual StrepTrap HP purification
Column:
Binding buffer:
Elution buffer:
Flow rate:
Two-step HisTrap HP and StrepTrap HP purification
Column:
Binding buffer:
Elution buffer:
Flow rate:
Column:
Sample:
Binding buffer:
Elution buffer:
Flow rate:
System:
SDS-PAGE
Acknowledgements:
Dual-tagged proteins
Products featured: StrepTrap HP, HisTrap HP, ÄKTAxpress, ExcelGel SDS Gradient 8–18, Multiphor II Electrophoresis System, LMW-SDS Marker Kit
Strep
Columns:
Sample: Strep E. coli
Binding buffer (StrepTrap HP):
Elution buffer (StrepTrap HP):
Binding buffer (HisTrap HP):
Elution buffer (HisTrap HP):
System:
Summary
0
100
200
300
400
500
600
700mAU
70 80 90 100 110 ml
A 280
StrepTrap HPelution
HisTrap HPelution
HisTrap HPflow through
66
30
Mr × 103
96
45
20.1
14.4
(His)6-mCherry-Strep-tag II
0
500
1000
1500
2000
mAU
70 80 90 100 110 120 130 140 ml
A 280
HisTrap HPelution
StrepTrap HPelution
Loopwash
StrepTrap HPflow through
Mr × 103
96
45
20.1
14.4
66
30(His)6-mCherry-Strep-tag II
imagination at work
Ela
nder
s i U
ppsa
la 2
008