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lac operon
DNA P lacI
C
E
O
lacZ
lacY
lacA
repressor CAP site pol-binding site operator structural genes
lactose
rep
T
terminator
DNA
9
lacZ gene - X-Gal chromogenic substrate
DNA P lacI
C
E
rep
O
lacZ
repressor CAP site pol-binding site operator
lacY
structural genes
IPTG
indoxyl
galactose
X-Gal
lacA
T
terminator
DNA
9
lacZ gene - X-Gal chromogenic substrate
DNS P lacI
C
E
rep
O
lacZ
lacY
repressor CAP site pol-binding site operator
pol
lacA
T
DNA
structural genes terminator
-gal
permease
transacetylase
lacZ gene in cloning
lacZ
DNA P lacI
C
E
O
-fragment
repressor CAP site pol-binding site operator
lacZ -fragment
lacZ-
lacA
structural genes


Foreign DNA
lacZ-
lacY
T
terminator
DNA
10
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7. Immunohistochemistry
- immunofluorescence
Fluorescent dye
A
Y
B
C
B
B
B
A
Y
Y
anti-C antibody
B
Cell surface antigens
Cytoplasmic antigenes
E
nucleus
D
cytoplasm
Direct method
Rabbit anti-C
Primary antibody
C
B
Cell surface antigenes
Citoplasmic antigenes
Goat anti-rabbit
Secondary antibody
E
nucleus
D
cytoplasm
Indirect method
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7. Immunohistochemistry
- immunoperoxidase-based methods
DAB + H2O2
Ni-DAB + H2O2
- peroxidase
Y
Y
- biotin – avidin – peroxidase
peroxidase
Anti-rabbit antibody
(secondary antibody)
B
A
Y
Y
peroxidase
Goat anti-rabbit
secondary antibody
Rabbit anti-C
primary antibody
anti-C antibody
C
B
B
Cell surfice antigens
cytoplazmic antigens
nucleus
D
cytoplasm
biotin
peroxidase
Y
Y
E
anti-rabbit antibody
(secondary antibody)
Indirect methods
anti-C antibody
peroxidase
peroxidase
avidin
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7. Immunohistochemistry
- immunoperoxidase-based methods
DAB + H2O2
Ni-DAB + H2O2
ABC method (ABC = avidin-biotin complex)
Y
Y
avidin
biotin
peroxidase
anti-rabbit antibody
(secondary antibody)
primary antibody
antigen
Y
Y
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7. Immunohistochemistry
alk. phosph.
- digoxigenin – anti-digoxigenin – alkaline phosphatase
B
A
Y
Y
Alkaline phosphatase
Y
Y
- alkaline phosphatase
Goat anti-rabbit
secondary antibody
Sheep anti-rabbit antibody
Rabbit anti-C antibody
Rabbit anti-C
primary antibody
C
B
B
Cell surfice antigens
cytoplasmic antigens
alk. phosph
E
cytoplasm
Indirect method
alk. phosph
DIG
anti-rabbit antibody
anti-C antitbody
Y
D
Y
Y
nucleus
DIG
anti-C antibody