THE BIOTECHNOLOGY
EDUCATION COMPANY
EDUCATION COMPANY
Troubleshooting Guide:
Protein
| PROBLEM: | CAUSE: | ANSWER: |
|---|---|---|
| Gel is not runninig properly | Running buffer was not properly prepared. | Check buffer protocol, make fresh buffer. |
| Wrong buffer used. | Check gel recipe, buffer must be compatible with the gel. | |
| Buffer volume is too low. | Buffer must fully cover the sample wells throughout the entire experiment. | |
| Gel is inserted in the wrong orientation. | Check with manufacturer for proper steup of the electrophoresis chamber. | |
| Malfunctioning electrophoresis chamber or power supply. | Consult with manufacturer of electrophoresis chamber or power supply. | |
| Tape at bottom of precast gel not removed. | Carefully remove tape before running the gel. | |
| Buffer volume is too low. | Buffer must fully cover the sample wells throughout the entire experiment. | |
| Electrodes not connected or polarity reversed. | Check electrode connections at the gel box and power supply. | |
| Poor band resolution or separation | Diffusion of samples before power was turned on. | Minimize time between loading samples and the start of electrophoresis. |
| The gel is old or expired. | Make fresh gels or order new pre-cast gels. | |
| Wrong concentration of acrylamide gel. | The kit is designed for 12% acrylamide gels, other concentrations will affect results. | |
| Smiling or frowning of bands | Proteins have been overloaded. | EDVOTEK® has optimized this kit to avoid overloading. Be sure to load the amount recommended by the protocol. |
| Wrong buffer used. | Check gel recipe, the buffer must be compatible with the gel. | |
| Incorrect voltage supplied to the gel. | Check the protocol for the recommended voltage. | |
| No bands on gel/smallest bands missing from gel | Proteins ran off gel. | Use the appropriate length of time for the chosen voltage. Be sure to monitor the tracking dye while the gel is running. For best results, the tracking dye should run 8-9 cm and should not be allowed to run off the gel. |
| Proteins have accumulated in the wells of the gel | Proteins have aggregated. | Ensure proteins have fully denatured; boil proteins for 5 minutes and load while still warm. |
| Bands are smeary and distorted | The gel has overheated. | Reduce voltage, check buffer concentration and dilute if necessary. |
| Bands are faint | Proteins have diffused or faded | Follow protocol for Protein InstaStain® to increase the contrast of protein bands. |
| Too little protein was loaded. | EDVOTEK® has optimized this kit to avoid underloading. Be sure to load the amount recommended by the protocol. |