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Document # 00522219-1009-00
Rockland, ME 04841 USAProSieve® ProTrack™ Dual Color Protein Loading Buffer
Instructions for Use
ProtocolIntroduction

Thaw Æ Mix Æ Denature Æ Load
1. Thaw the Reducing Agent at room temperature.
Dissolve precipitated solids in the Loading buffer(if any) at 37°C.
2. Gently mix the loading buffer and reducing agent
to ensure that the solutions are homogeneous.
3. Add appropriate 20X Reducing Agent,ProSieve® ProTrack™ Loading Buffer andprotein sample solution (protein sample in water)
into clean microcentrifuge tube. Refer to tablebelow for recommended volumes.
4. Heat samples at 100°C for 3-5 minutes.
5. Centrifuge briefly and apply directly to a SDSpolyacrylamide gel.

The ProSieve® ProTrack™ Dual Color Protein
Loading Buffer protects proteins from heatdegradation during the sample preparation step, as
well against pH changes during the SDS-PAGE run.
Some proteins are sensitive to pH changes that
result from temperature fluctuations during
electrophoresis in Tris buffers. The optimizedcomposition of the loading buffer prevents protein
degradation during sample heating prior to SDSPAGE1 as well as during the electrophoresis run.
The loading buffer contains two tracking dyes: blue(bromophenol blue) for tracking the progress ofelectrophoresis and pink (pyronin Y) for monitoringof protein transfer to the membrane during Western
blotting procedures. ProSieve® ProTrack™ Loading
Buffer also contains SDS and DTT for complete
disruption of all high-order protein structures.

Recommended Volumes for Gel Loading

Gel Loading
Volume14 μl
20 μl
30 μl
40 μlProtein samples prepared withProSieve® ProTrack™ Dual Color Protein Loading Buffer

Gel

DTT*0.7 μl
1 μl
1.5 μl2 μlLoadingDye
3.5 μl5 μl
7.5 μl10 μl

SampleSolution†
9.8 μl
14 μl
21 μl
28 μl

* For silver staining see Note.
†diH2O may need to be added to samples to yield
volumeBlotNoteContents•

Cat. No. 00193861
4X ProSieve® ProTrack™ Loading Buffer, 5 ml
(1000 applications @ 20ul)
Storage Buffer
0.25 M Tris-HCl, 1.6 mM EDTA (pH 8.5 at 25°C), 8%(w/v) SDS, 40% (w/v) glycerol, 0.04% (w/v)
bromophenol blue, 0.024% (w/v) pyronin Y.

••

Store at Room Temperature
20X Reducing Agent (2 M DTT), 1 ml
Store at -20°C
All trademarks herein are marks of Lonza Groupor its subsidiaries.

1For silver staining, DTT concentration in thesample should not exceed 50 mM. Higher DTT
concentration in protein sample may cause
streaking or yellowing of the gel. If initial protein
sample contains >50 mM DTT, do not add
Reducing Agent into the protein loading buffer.
Due to presence of SDS, the loading buffer is
not suitable for native polyacrylamide gelelectrophoresis.
Generally pyronin Y migrates faster thanbromophenol blue. In higher percentage SDSpolyacrylamide gels (for example 15%) pyronin
Y dye migrates slower than bromophenol blue.

Reference
1. Electrophoresis in practice, 4th edition,
Westermeier, R., Wiley-VCH, 2005.
Product Safety
For details regarding product safety, see Material Safety
Data Sheet (MSDS); call +1 (800) 638-8174 for extra
copies of the MSDS. Emergency after hours, call collect+1 (303) 595-9048.

Manufactured for Lonza Rockland, Inc.
For Research Use Only.
Not For Use In Diagnostic Procedures.
©2009 Lonza Rockland, Inc.
All rights reserved.

All trademarks herein are marks of Lonza Groupor its subsidiaries.

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