User Manual

Section 8
Solutions for Protein Applications
8.1 Solutions for Nitrocellulose Membrane
Tris Buffered Saline, 1x TBS, 2 L
20 mM Tris-HCl, pH 7.5
500 mM NaCl
Dissolve 4.84 g Tris, 58.48 g NaCl in ~1.5 L distilled, deionized H
2
O. Adjust to pH 7.5
with HCl. Adjust the volume to 2 L with ddH
2
O.
Tween 20 Wash Solution, 1x TTBS, 1 L
20 mM Tris, pH 7.5
500 mM NaCl
0.05% Tween 20
Add 0.5 ml Tween 20 to 1 L of TBS.
Blocking Solution, 100 ml
1% BSA-TBS
Add 1.0 g bovine serum albumin (BSA) to 100 ml TBS. Stir to dissolve.
Antibody Buffer, 200 ml
1% BSA-TTBS
Add 2 g BSA to 200 ml TTBS. Stir to dissolve. 100 ml should be reserved for primary
antibody and an equal volume for dilution of the secondary antibody conjugate.
Primary Antibody Solution, 100 ml
Dilute antigen specific primary antibody to the appropriate titer in 100 ml of antibody
buffer.
Secondary Antibody Solution, 100 ml
Dilute species-specific Bio-Rad secondary antibody conjugate, 1:3,000 by adding 33 µl
of conjugate to 100 ml of antibody buffer.
Color Development Solution
The specific chemicals and buffers are dependent on the enzyme conjugate being
used. See the Immun-Blot
®
assay kit instruction manual for details on how to make the
appropriate solution.
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