TruPAGE™ Troubleshooting Guide

Problem Possible Cause Suggested Solution
Inner (Cathode) chamber The gel is not properly seated in the chamber. Ensure the gel cassette is seated in the chamber according to the gel running equipment manufacturer's instructions.
10 × 8 cm gel is being used with equipment designed for 10 × 10 cm gels. 10 × 8 cm gel cassettes are 2.5 mm thinner than 10 × 10 cm cassettes and cannot be used with gel boxes designed for 10 × 10 cm gels. Always use gels compatible with equipment available. Equipment compatibility information is available on TruPAGE gel product detail pages and Technical Bulletin online.
Bio-Rad running box gasket was not flipped to the flat side. Flip the device gasket to its flat backside. Instructions can be found online in the TruPAGE Technical Bulletin.
Gel does not fit into the running box chamber. 10 × 10 cm gel is being used with equipment designed for 10 × 8 cm gels. Only use 10 × 8 cm and 10 × 10 cm gels with gel running equipment designed for gels of these formats. Equipment compatibility information is available on TruPAGE gel product detail pages and online in the TruPAGE Technical Bulletin.
No current during run. Buffer leaking from the inner chamber. See Problem, “Inner (Cathode) chamber buffer leaking” for resolution.
Low voltage output from the power supply. Ensure that running voltage is set according to instructions found in the TruPAGE Quick Reference Guide provided in every box of gels. Ensure that the power supply is functioning properly.
Running buffer was not properly diluted. Make fresh 1× TruPAGE Running Buffer from 20× concentrated solution. Buffer formulations are provided in the Quick Reference Guide in every box of gels and online TruPAGE Technical Bulletin.
Gels overheating during electrophoresis. Recommended run voltage exceeded. Follow TruPAGE gel running instructions found in the Quick Reference Guide provided in every box of gels and online in the TruPAGE Technical Bulletin.
Outer (anode) buffer chamber is not filled properly. Ensure to fill the outer chamber, as the buffer in the outer chamber serves as coolant during gel run.
TruPAGE gels are being run alongside gels from other manufacturers. Gels with differing gel chemistry have different electrical resistivity properties. Do not run gels from different manufacturers alongside each other in the same gel running box.
Do not run TruPAGE and another type of gel (i.e. Tris-Glycine) in different tanks but on the same power supply. The TruPAGE gel and buffer system pulls more current from the power supply increasing the temperature in the TruPAGE tank which results in smiling.
Dye front changes color (i.e., yellow). Underfilled outer (anode) buffer chamber is causing buffer starvation. Add more 1× TruPAGE Running Buffer to the outer buffer chamber.
Misshaped protein bands and/or crooked lanes. Gel overheating during run. See Problem “Gels overheating during electrophoresis” for resolution.
Incompatible running buffer used. Only run TruPAGE gels using TruPAGE formulated running and sample loading buffers. Buffer compositions can be found online in the TruPAGE Technical Bulletin and in the Quick Reference Guide provided in every box of gels.
Excessive streaking of protein bands. Incompatible sample buffer used. Always prepare samples with TruPAGE LDS Sample Buffer. Buffer composition can be found in the online TruPAGE Technical Bulletin and Quick Reference Guide provided in every box of gels.
Too much protein loaded into the wells. Prevent sample overload by limiting the amount of protein loaded per well. Maximum recommended protein loading: 10 μg for purified protein, 100 μg for lysate.
Precipitate/particulates in the sample. Centrifuge the sample to remove any insoluble matter or decrease total sample concentration to ensure complete solubility of proteins.
Salt concentration is too high. If sample lysis/storage buffer contains a high salt concentration, it may be necessary to perform dialysis prior to starting sample preparation for electrophoresis.
Protein samples diffusing rapidly throughout the gel. Gel has been frozen. Always store TruPAGE gels at recommended storage conditions (2–8 °C).
Prestained protein markers indicate different apparent molecular masses. Prestained protein markers used have not been calibrated for use with TruPAGE gels. Dyes used for staining protein markers behave differently in different buffer systems, which will affect the migration patterns of prestained proteins. Calibrate prestained protein markers against unstained proteins of known size or use ColorBurst™ Electrophoresis Marker (Catalog Number C1992), a solution of precalibrated, prestained protein markers.