Bio-Rad Biotinylated Standards - Manual (Page 4)

Section 2
Specifications
Table 1 Composition of the Biotinylated SDS-PAGE
Standards
MW
Protein Source (dalton) High Low Broad Reference
Myosin rabbit skeletal 200,000 x x 2
muscle
ß-galactosidase E. coli 116,250 x x 3
Phosphorylase B rabbit muscle 97,400 x x x 4
Serum albumin bovine 66,200 x x x 5
Ovalbumin hen egg white 45,000 x x x 6
Carbonic bovine 31,000 x x 7
anhydrase
Trypsin inhibitor soybean 21,500 x x 8
Lysozyme hen egg white 14,400 x x 9
Aprotinin bovine pancreas 6,500 x 10
Kit Contents
Biotin. Stds. Biotin. Stds. Biotin. Stds.
Low Range* High Range* Broad Range Avidin-HRP † Avidin-AP ††
161-0307 ••• •••
161-0308 ••• •••
161-0312 ••• •••
161-0313 ••• •••
161-0321 ••• •••
161-0322 ••• •••
† * The biotinylated standards contain approximately 130 µ g total protein in
50% glycerol, 150 mM NaCl, 100 mM DTT, 3 mM NaN
3
.
† † Avidin-horseradish peroxidase conjugate is supplied in 10 mM NaPO
4
, 150
mM NaCl, pH 7.2, containing 1.0% BSA as a stabilizer and 0.01%
Thimerosal as a bacteriostat.
†† Avidin-alkaline phosphatase is supplied in 10 mM Tris pH 8.0, 150 mM
NaCl, 1 mM MgCl
2
, containing 1.0% BSA as a stabilizer and 0.1% sodium
azide as a bacteriostat.
3
A B
Fig. 1. 12% SDS-polyacrylamide gels run by the method of
Laemmli in the Mini-PROTEAN ® ll cell, and transferred to nitro-
cellulose in the Mini Trans-Blot ® cell. Both gels had the same
loading pattern: Lanes 1 and 6, total human serum; Lanes 2 and 7,
E. coli lysate; Lanes 3 and 8, human transferrin; Lanes 4 and 9,
Biotinylated SDS-PAGE Standards, Low Range; Lane 5, SDS-PAGE
Molecular Weight Standards, Low Range.
A) Blot was stained with Bio-Rad's Colloidal Gold Total Protein Stain.
The biotinylated standards in lane 4 show minimal mobility changes
compared to the untreated standards in lane 5.
B) Blot was treated with rabbit anti-human transferrin as the primary
antibody. The second antibody solution contained Avidin-HRP and
GAR-HRP. The color reaction was done using HRP Color
Development Reagent. The blot shows the human transferrin band
was positively identified in lanes 6 and 8. The other proteins were
not detected by the Avidin-HRP, showing that Avidin-HRP binds
specifically to the biotinylated standards.
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