DNA Methylation EIA Short Protocol
1. EIA Buffer - Dilute with 90 ml of UltraPure water.
2. Wash Buffer - Dilute the 5 ml vial to 2 L and add
1 ml Tween 20 or dilute the 12.5 ml vial to 5 L and
add 2.5 ml Tween 20.
3. Tracer - Reconstitute the 100 dtn vial with 6 ml of EIA
Buffer or the 500 dtn vial with 30 ml EIA Buffer.
4. Antibody - Reconsitute the 100 dtn vial with 6 ml of
EIA Buffer or the 500 dtn vial with 30 ml EIA Buffer.
5. Standard - Prepare as described in the figure below.
100 μl
500 μl
500 μl
100 μl
S1
S2
500 μl
S3
500 μl
500 μl
S4
S5
500 μl
S6
500 μl
S7
S8
Final
900 μl
EIA
Buffer
900 μl
UltraPure
H2 O
15 μg/ml
Standard
1.5 μg/ml
Bulk Standard
Steps
Reagent
1. Add
Reagents
EIA Buffer
Standard/Sample
Tracer
Antibody
2. Incubate
75
ng/ml
150
ng/ml
Blank
5. Incubate
6. Read
500 μl
EIA
Buffer
37.5
ng/ml
TA
500 μl
EIA
Buffer
18.75
ng/ml
500 μl
EIA
Buffer
9.38
ng/ml
NSB
500 μl
EIA
Buffer
4.69
ng/ml
B0
100 μl
50 μl
------50 μl
50 μl
--50 μl
---Cover plate and incubate 18 hours at 4°C
3. Wash
4. Add
Reagents
500 μl
EIA
Buffer
500 μl
EIA
Buffer
2.34
ng/ml
500 μl
EIA
Buffer
1.17
ng/ml
Std/Sample
-50 μl
50 μl
50 μl
Wash all wells five times
Tracer
--
5 μl
--
--
--
Ellman's
200 μl
200 μl
200 μl
200 μl
200 μl
Cover plate and incubate 90-120 minutes RT with gentle shaking
Read plate at a wavelength between 405-420 nm
A
B
C
D
E
F
G
H
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