Related Experiment Video
Updated: May 22, 2026
06:58
Real-time Tracking of DNA Fragment Separation by Smartphone
Published on: June 1, 2017
Common artifacts and mistakes made in electrophoresis
Biji T Kurien1, R Hal Scofield
1Department of Veterans Affairs, University of Oklahoma Health Sciences Center, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA. biji-kurien@omrf.org
Methods in Molecular Biology (Clifton, N.J.)
|May 16, 2012
Summary
Subtle experimental artifacts and common researcher errors can compromise protein analysis results. Careful attention to sample preparation, buffer conditions, and electrophoresis techniques is crucial for accurate scientific findings.
Area of Science:
- Biochemistry
- Proteomics
- Molecular Biology
Background:
- Experimental artifacts can significantly impact the reliability of protein analysis.
- Common laboratory practices may introduce subtle errors affecting experimental outcomes.
Purpose of the Study:
- To identify and detail common artifacts and researcher-induced errors in protein experimentation.
- To provide guidance on avoiding pitfalls in experimental design and execution.
Main Methods:
- Review of potential sources of experimental artifacts in protein sample preparation and electrophoresis.
- Analysis of common procedural mistakes made by researchers during electrophoresis and sample preparation.
Main Results:
- Identified artifacts include protease activity, Asp-Pro bond cleavage, keratin contamination, chemical leaching, and urea impurities.
- Common errors involve gel cross-linking calculation, polymerization, sample aggregation, buffer titration, sample preparation, protein loading, buffer ratios, incomplete buffer removal, and isoelectric focusing.
Conclusions:
- Addressing these artifacts and researcher errors is essential for generating reproducible and accurate experimental results.
- Meticulous attention to detail in all experimental stages is critical for successful protein analysis.
Related Concept Videos
01:20Electrophoresis: Overview
Electrophoresis is a powerful analytical separation technique that relies on the differential migration of charged species when subjected to an electric field. The core strength of electrophoresis lies in its ability to separate high-molecular-weight species in complex mixtures. It has found widespread use in biochemistry, molecular biology, and analytical chemistry, allowing the separation of compounds like amino acids, nucleotides, carbohydrates, and proteins with excellent resolution.
There...
There...
01:22Two-dimensional Gel Electrophoresis
Two-dimensional gel electrophoresis is a high-resolution protein separation method first introduced by O' Farrell and Klose in 1975. This method involves protein separation by two dimensions, mass and charge, making it more accurate than one-dimensional gel electrophoresis.
The first dimension separation uses the isoelectric focusing or IEF technique performed on immobilized pH gradient (IPG) strips that separate proteins according to their isoelectric points.
Biological samples, such as cells...
The first dimension separation uses the isoelectric focusing or IEF technique performed on immobilized pH gradient (IPG) strips that separate proteins according to their isoelectric points.
Biological samples, such as cells...
01:20Capillary Electrophoresis: Instrumentation
Capillary electrophoresis instrumentation typically consists of several key components. A high-voltage power supply generates the electric field necessary for the separation by connecting to an anode (the positively charged electrode) and a cathode (the negatively charged electrode) located in buffer reservoirs at each end of the capillary tube. The system includes a sample vial, a fused silica capillary tube coated with polyimide for mechanical strength through which the sample components...
02:35DNA Agarose Gel Electrophoresis
Agarose gel electrophoresis is a laboratory technique commonly used to separate DNA fragments by size. However, it can also be used to isolate and purify DNA fragments using a gel extraction protocol.
Gel extraction follows five major steps: running gel electrophoresis to separate fragments, isolating the individual bands, extracting DNA from those bands, and removing the dye and salts from the extracted mixture to obtain pure DNA.
In cloning experiments, both the insert and vector DNA...
Gel extraction follows five major steps: running gel electrophoresis to separate fragments, isolating the individual bands, extracting DNA from those bands, and removing the dye and salts from the extracted mixture to obtain pure DNA.
In cloning experiments, both the insert and vector DNA...
01:30Capillary Electrophoresis: Applications
Capillary electrophoretic separations offer various modes, each with unique applications. These modes include capillary zone electrophoresis, capillary gel electrophoresis, capillary array electrophoresis, capillary isoelectric focusing, capillary isotachophoresis, micellar electrokinetic chromatography, and capillary electrochromatography.
Capillary zone electrophoresis (CZE) separates ionic components based on their electrophoretic mobility. It has been used to separate proteins, amino acids,...
Capillary zone electrophoresis (CZE) separates ionic components based on their electrophoretic mobility. It has been used to separate proteins, amino acids,...
02:57Southern Blot
Agarose gel electrophoresis is very useful in separating DNA fragments by size. Running a DNA ladder containing fragments of the known length alongside the sample helps determine the approximate length of the sample DNA fragments. However, additional steps are needed to verify the sequence identity of the sample DNA fragments.
Denatured DNA fragments must be transferred onto a carrier membrane from the gel to make it accessible to a probe - a small ssDNA fragment complementary to the target DNA...
Denatured DNA fragments must be transferred onto a carrier membrane from the gel to make it accessible to a probe - a small ssDNA fragment complementary to the target DNA...